The validity of the model must be tested. This is usually done by splitting the sample set into two sets; one set for calibration and the other for validation. If there are not enough samples, "leave-one-out" cross validation can be performed. This means leaving one sample out, using the rest of the samples to build a calibration model and then using the model to predict the one left out. The advantage of doing cross validation is that unlike calibration with a full data set, the sample being predicted is not included in the calibration model. Thus, the model can be tested independently.

#### **2.5 Prediction**

Finally, the calibration can be used to predict future unknowns, assuming the unknowns are in the same sample population as those used in the calibration set. Whether the unknown is an outlier needs to be tested.

#### **2.6 Summary**

Applying an NIR analyzer to a particular application requires the development of a reliable calibration model. The most important steps involve a thorough consideration of experimental design and multivariate calibration. Once this is established, one can enjoy the advantages of the NIR analysis. The speed of the analysis will save time and avoid mistakes instantaneously. The speed advantage is so valuable to engineers involved with on-line process monitoring that instruments are routinely installed in or near the process line with feedback loops. With an NIR analyzer such as QualitySpec® Pro spectrometer, samples can be non-invasively analyzed on-the-spot, dramatically reducing costly and time consuming laboratory analysis as well as preventing unnecessary product waste and/or downtime. The low absorptivity in the NIR region allows measurements to be taken on raw materials, in process and finshed product without elaborate sample preparation. In the food, agricultural, pharmaceutical, polymer, cosmetics, environmental, textile, and medical fields, NIR analysis serves a wide range of applications, with still many unknown applications waiting to be discovered. With the maturity of this technique, more and more people will use NIR analysis for convenience and flexibility.

#### **2.7 Wavenumber and wavelength**

where

Y = the number of nanometers (nm) X = the number of wavenumbers (cm-1)

$$\text{Y} = 10^{\text{v}} / \text{ 28571} = \text{350}$$

Y = 107 / X

Resolution in cm-1, ( 1 cm R − ) is dependent upon wavelength position. So, Resolution in nanometers, (Rnm) is calculated as follows:

$$\begin{array}{lcl} \mathbf{R}\_{\text{nm}} &= +/ \text{ - [Y - Y']} \\ &= +/ \text{ - } \{ [10^{\top}/\text{\AA}] \cdot [10^{\top}/\text{\AA}] \} \\ &= +/ \text{ - } 10^{\top} \text{ \* [[1/\text{\AA}] - [1/\text{\AA}]] } \\ &= +/ \text{ - } 10^{\top} \text{ \* [[1/\text{\AA}] - [1/\text{\AA} - \text{R}\_{\text{cm}} \text{-}]] } \} \end{array}$$

**Part 3**

**Quality Control in Clinics**

#### **3. References**

**Part 3**

**Quality Control in Clinics**

248 Wide Spectra of Quality Control

[1] Burns D. A. and E. W. Ciurczak (Eds.), Handbook of Near-Infrared Analysis, (Volume 13 in Practical Spectroscopy Series), Marcel Dekker, Inc., New York, 1992 [2] Hildrum K. I., T. Isaksson, T. Naes and A. Tandberg (Eds.), Near Infra-red Spectroscopy,

[3] Murray I. and I. A. Cowe (Eds.), Making Light Work: Advances in Near Infrared

[4] Burns D. A. and E. W. Ciurczak (Eds.), Handbook of Near-Infrared Analysis, (Volume 13

(Ellis Horwood Series in Analytical Chemistry), Ellis Horwood, Ltd., England, 1992

Spectroscopy, 4th International Conference on Near Infrared Spectroscopy, Aberdeen, Scotland, August 19-23, 1991, Weinheim, New York, Basel, Cambridge,

in Practical Spectroscopy Series), Marcel Dekker, Inc., New York, 1992, pgs 393-395.

**3. References**

VCH, 1992

**14**

**Quality Control in Hospital Bone Banking**

The use of allogenic bone transplantation is nowadays a standard orthopaedic procedure. It is widely used for reconstruction of bone defects that arise from trauma (Friedlaender 1987), infection, resection of bone tumours (Mankin et al. 1996) or it is used in spinal fusion (Raizman et al. 2009) and as impaction grafting in revision of total joint arthroplasty (Slooff et al. 1996). Although autologous bone is generally preferred because of its osteoconductive and osteoinductive activity, autologous bone is often not sufficiently available and comes with donor site morbidity (Summers & Eisenstein 1989). Therefore allogenic bone grafts are often used in orthopaedic procedures. These bone allografts are provided by an orthopaedic bone bank. It might be financially attractive for a hospital to manage its own local hospital bone bank, especially if they perform many procedures in which bone allograft is used. The main advantage of managing a hospital bone bank however, is the easy accessibility to and availability of bone allograft. The bone allografts in a hospital bone bank are femoral heads obtained from suitable patients who underwent total hip replacement surgery. Management of an orthopaedic bone bank is a complex process. The bone bank procedure has to meet the requirements of the national law and European guidelines 2004/23/EC and 2006/86/EC. This law states the technical requirements for coding, processing, preserving, storing, and distributing of human tissue and cells. Human tissue should be traceable and serious side effects and incidents with human tissue and cells should be reported. The bone bank procedure should be carefully described in an extensive protocol. Neither in the Netherlands, nor in any other European country, there are official guidelines for the organization and management of an orthopaedic bone bank. Our bone banking procedure protocol is based on guidelines of The American Association of Tissue Banks (AATB 1993), the criteria of the Council for Blood Transfusion of the Netherlands Red Cross (Richtlijn Bloedtransfusie 2004), the recently merged Netherlands Bone Bank Foundation (NBF) and Bio Implant Services (BIS); (NBF-BIS Foundation 2010) and the guidelines of the European Association of Musculoskeletal Transplantation (EAMST). The latter has been discontinued because of diverging European legislation. This bone bank protocol extensively describes the procedure, which includes a thorough questionnaire for donor selection, extensive serological, bacteriological and histopathological examination, as well as standard procedures for registration, processing, preservation, storage and distribution of bone allografts (Zwitser et al. 2010). In this chapter we describe our local hospital bone banking

**1. Introduction**

Eline W. Zwitser and Barend J. van Royen

*Department of Orthopaedic Surgery VU Medical Center Amsterdam*

*The Netherlands*

### **Quality Control in Hospital Bone Banking**

Eline W. Zwitser and Barend J. van Royen

*Department of Orthopaedic Surgery VU Medical Center Amsterdam The Netherlands*

#### **1. Introduction**

The use of allogenic bone transplantation is nowadays a standard orthopaedic procedure. It is widely used for reconstruction of bone defects that arise from trauma (Friedlaender 1987), infection, resection of bone tumours (Mankin et al. 1996) or it is used in spinal fusion (Raizman et al. 2009) and as impaction grafting in revision of total joint arthroplasty (Slooff et al. 1996). Although autologous bone is generally preferred because of its osteoconductive and osteoinductive activity, autologous bone is often not sufficiently available and comes with donor site morbidity (Summers & Eisenstein 1989). Therefore allogenic bone grafts are often used in orthopaedic procedures. These bone allografts are provided by an orthopaedic bone bank. It might be financially attractive for a hospital to manage its own local hospital bone bank, especially if they perform many procedures in which bone allograft is used. The main advantage of managing a hospital bone bank however, is the easy accessibility to and availability of bone allograft. The bone allografts in a hospital bone bank are femoral heads obtained from suitable patients who underwent total hip replacement surgery. Management of an orthopaedic bone bank is a complex process. The bone bank procedure has to meet the requirements of the national law and European guidelines 2004/23/EC and 2006/86/EC. This law states the technical requirements for coding, processing, preserving, storing, and distributing of human tissue and cells. Human tissue should be traceable and serious side effects and incidents with human tissue and cells should be reported. The bone bank procedure should be carefully described in an extensive protocol. Neither in the Netherlands, nor in any other European country, there are official guidelines for the organization and management of an orthopaedic bone bank. Our bone banking procedure protocol is based on guidelines of The American Association of Tissue Banks (AATB 1993), the criteria of the Council for Blood Transfusion of the Netherlands Red Cross (Richtlijn Bloedtransfusie 2004), the recently merged Netherlands Bone Bank Foundation (NBF) and Bio Implant Services (BIS); (NBF-BIS Foundation 2010) and the guidelines of the European Association of Musculoskeletal Transplantation (EAMST). The latter has been discontinued because of diverging European legislation. This bone bank protocol extensively describes the procedure, which includes a thorough questionnaire for donor selection, extensive serological, bacteriological and histopathological examination, as well as standard procedures for registration, processing, preservation, storage and distribution of bone allografts (Zwitser et al. 2010). In this chapter we describe our local hospital bone banking

Quality Control in Hospital Bone Banking 253

Only fresh amputated limbs could be used as a donor allograft. For this reason, autografts were used much more frequently than allografts. In the 1940's storage methods were developed for preservation longer than a few days or hours by refrigeration or freezing. In 1949 in Bethesda, Maryland the first United States Navy Tissue Bank was established, because of a military need for bone allografts (Hyatt 1950). In that Tissue Bank allografts were obtained from the nearby National Naval Medical Centre. In addition, they developed a method for freeze-drying of allografts by lyophilisation, which made it possible to store and preserve allografts for several years without the need for refrigeration or freezing (Kreuz et al. 1951). In the first century of bone transplantation disease transmission was not of great concern. In addition, serological tests for transmittable diseases other than syphilis were not available. The first case ever of the transmission of viral disease by frozen bone was reported by Shutkin in 1954. The donor had undergone an above the knee amputation. The allograft bone was cut into portions under aseptic conditions, placed in double sterile containers and frozen at a temperature of -10 to -20 °C. Five months later, the bone was implanted into a medical student and transmitted hepatitis B. In the early 1980's the first publications concerning a new disease AIDS were published (Centers for Disease Control [CDC] 1981, 1982). Only a few years later in 1984 the first transmission of disease by bone allograft was reported (CDC 1988). A serological test was not yet available at the time of transmission. Even with the first serological tests used for screening purposes another transmission occurred in 1985, due to a very recent donor infection in the so-called "window period" of testing, with a less accurate test. Therefore, in an expert conference guidelines and recommendations were developed (CDC 1988). The most important conclusion drawn was that the disease was transmitted by blood and bone marrow containing allografts. Recommendations concerned donor screening, testing and re-testing of living donors after 6 months. The constant update for screening of donors for infectious diseases proved to be important in the subsequent years with two reports on transmission of hepatitis C virus by bone allografts (Eggen & Nordbø 1992; Conrad et al 1995). In the following years tissue banks developed better techniques for processing and preparation of bone allografts and more reliable blood tests came to market (Busch 1991, 1994; Alter et al. 1990). However, donor screening methods are constantly updated and revised with the introduction of new infectious diseases, like SARS (Lam et al. 2004). The safety of allograft bone transplants can

never be taken for granted, but recent safety records for bone allografts are excellent.

(safe) bone allografts has increased (Nielsen et al. 2001).

**3. Indications for the use of bone allograft**

As more complex orthopaedic surgical procedures are performed nowadays the need for

Massive bone defects can arise from trauma, infection, osteolysis after arthroplasty or resection of bone tumours and are a challenging problem in orthopaedic practice. These bone defects can be filled with either autograft or allograft bone transplants. Ideally, autograft is preferred because of its osteoconductive and osteoinductive activity. However, autografts are available in limited number and size and therefore not sufficiently available. In addition harvesting is associated with extended surgical time and involves donor site morbidity (Aro&Aho 1993; Summers and Eisenstein 1989). Therefore allografts supplied by a bone bank are commonly used instead. Allogenic bone exclusively has osteoconductive activity; it serves as an acellular mineralized frame against which newly formed bone gets deposited (Elves and Pratt 1975; Urist 1953). Indications for the use of allografts are wide

procedure and protocol. This constantly updated protocol is of the utmost importance in order to prevent the transmission of infectious diseases. Because of the potential risk of transmission of diseases from donor to recipient we performed routine histological examination in the screening protocol. We found a relatively high percentage of pathological conditions in retrieved femoral heads (Zwitser et al. 2009; Sugihara et al. 1999). Therefore, we recommend the routine histopathological evaluation of all femoral heads removed during elective total hip arthroplasty as a tool for quality control. The costperformance ratio of routine histopathological evaluation is discussed in literature (Kocher et al. 2000; Meding et al. 2000; Lawrence et al. 1999; Campbell et al. 1997). Therefore we performed an intern evaluation of the bone banking process. We compared the costs made to harvest, store and implant one bone allograft of our bone bank and the costs of one allograft obtained from the central bone bank. Furthermore this evaluation brought valuable information of improvements to be made. We describe these conclusions and have suggestions how to further improve the quality and cost effectiveness of the bone banking process in the near future.

#### **2. History of bone transplantation**

The history of bone transplantation can be traced back to the seventeenth century. In 1668 Job van Meekeren, a Dutch surgeon was the first to perform bone transplantation (Schweiberer et al. 1990). He repaired a skull defect in a soldier with part of a skull from a dog. As soon as the soldier was informed about the transplant, he requested immediate removal of the dog's skull. This was not possible because the xenograft was already fully incorporated in the man's skull. The first human allograft ever reported describes a case of a bone transplant in a young male who suffered an osteomyelitis of the entire humeral shaft (MacEwen 1881). In 1881, the treatment for osteomyelitis was surgical debridement or resection of the affected bone. After this surgical procedure, it took several years for the infection to extinguish, where after the surgeon could replace the bony defect with fresh allografts from diaphysis of tibial shaft. In the following seven years these allografts slowly but successfully incorporated in the recipient humeral shaft (MacEwen 1909). In the following decades the technique of transplantation of large allografts in septic arthritis and osteomyelitis was further developed and popularised by a German surgeon. He used fresh long bones of amputated limbs and used them as osteoarticular allografts with a reported success rate of 50% (Lexer 1908, 1925). Later, in 1929 Alexander Fleming discovered the antibacterial properties of penicillium and treatment modalities of osteomyelitis changed (Fleming 1929). In the subsequent years antibiotics were further developed and introduced for clinical medical use in the 1950's. From now on, the treatment of choice for osteomyelitis consisted of appliance of antibiotics in stead of surgery.

Transplantation of large allografts was applied as a limb-saving treatment in high grade malignant bone tumours of the lower extremity (Parrish 1973, Mankin et al. 1976). In a series of 19 allograft replacements for osseous malignancies satisfactory results were reported in 75% of patients. In the largest series of two hundred lower extremity osteoarticular allografts performed between 1976 and 1997 for malignant bone tumours, results were diminished by radiation and chemotherapy (Hazan et al. 2001).

In the first century of bone transplantation the greatest impediment to the use of allografts was availability of fresh bone grafts, because there were no means for preservation.

procedure and protocol. This constantly updated protocol is of the utmost importance in order to prevent the transmission of infectious diseases. Because of the potential risk of transmission of diseases from donor to recipient we performed routine histological examination in the screening protocol. We found a relatively high percentage of pathological conditions in retrieved femoral heads (Zwitser et al. 2009; Sugihara et al. 1999). Therefore, we recommend the routine histopathological evaluation of all femoral heads removed during elective total hip arthroplasty as a tool for quality control. The costperformance ratio of routine histopathological evaluation is discussed in literature (Kocher et al. 2000; Meding et al. 2000; Lawrence et al. 1999; Campbell et al. 1997). Therefore we performed an intern evaluation of the bone banking process. We compared the costs made to harvest, store and implant one bone allograft of our bone bank and the costs of one allograft obtained from the central bone bank. Furthermore this evaluation brought valuable information of improvements to be made. We describe these conclusions and have suggestions how to further improve the quality and cost effectiveness of the bone banking

The history of bone transplantation can be traced back to the seventeenth century. In 1668 Job van Meekeren, a Dutch surgeon was the first to perform bone transplantation (Schweiberer et al. 1990). He repaired a skull defect in a soldier with part of a skull from a dog. As soon as the soldier was informed about the transplant, he requested immediate removal of the dog's skull. This was not possible because the xenograft was already fully incorporated in the man's skull. The first human allograft ever reported describes a case of a bone transplant in a young male who suffered an osteomyelitis of the entire humeral shaft (MacEwen 1881). In 1881, the treatment for osteomyelitis was surgical debridement or resection of the affected bone. After this surgical procedure, it took several years for the infection to extinguish, where after the surgeon could replace the bony defect with fresh allografts from diaphysis of tibial shaft. In the following seven years these allografts slowly but successfully incorporated in the recipient humeral shaft (MacEwen 1909). In the following decades the technique of transplantation of large allografts in septic arthritis and osteomyelitis was further developed and popularised by a German surgeon. He used fresh long bones of amputated limbs and used them as osteoarticular allografts with a reported success rate of 50% (Lexer 1908, 1925). Later, in 1929 Alexander Fleming discovered the antibacterial properties of penicillium and treatment modalities of osteomyelitis changed (Fleming 1929). In the subsequent years antibiotics were further developed and introduced for clinical medical use in the 1950's. From now on, the treatment of choice for osteomyelitis

Transplantation of large allografts was applied as a limb-saving treatment in high grade malignant bone tumours of the lower extremity (Parrish 1973, Mankin et al. 1976). In a series of 19 allograft replacements for osseous malignancies satisfactory results were reported in 75% of patients. In the largest series of two hundred lower extremity osteoarticular allografts performed between 1976 and 1997 for malignant bone tumours, results were

In the first century of bone transplantation the greatest impediment to the use of allografts

was availability of fresh bone grafts, because there were no means for preservation.

process in the near future.

**2. History of bone transplantation**

consisted of appliance of antibiotics in stead of surgery.

diminished by radiation and chemotherapy (Hazan et al. 2001).

Only fresh amputated limbs could be used as a donor allograft. For this reason, autografts were used much more frequently than allografts. In the 1940's storage methods were developed for preservation longer than a few days or hours by refrigeration or freezing.

 In 1949 in Bethesda, Maryland the first United States Navy Tissue Bank was established, because of a military need for bone allografts (Hyatt 1950). In that Tissue Bank allografts were obtained from the nearby National Naval Medical Centre. In addition, they developed a method for freeze-drying of allografts by lyophilisation, which made it possible to store and preserve allografts for several years without the need for refrigeration or freezing (Kreuz et al. 1951). In the first century of bone transplantation disease transmission was not of great concern. In addition, serological tests for transmittable diseases other than syphilis were not available. The first case ever of the transmission of viral disease by frozen bone was reported by Shutkin in 1954. The donor had undergone an above the knee amputation. The allograft bone was cut into portions under aseptic conditions, placed in double sterile containers and frozen at a temperature of -10 to -20 °C. Five months later, the bone was implanted into a medical student and transmitted hepatitis B. In the early 1980's the first publications concerning a new disease AIDS were published (Centers for Disease Control [CDC] 1981, 1982). Only a few years later in 1984 the first transmission of disease by bone allograft was reported (CDC 1988). A serological test was not yet available at the time of transmission. Even with the first serological tests used for screening purposes another transmission occurred in 1985, due to a very recent donor infection in the so-called "window period" of testing, with a less accurate test. Therefore, in an expert conference guidelines and recommendations were developed (CDC 1988). The most important conclusion drawn was that the disease was transmitted by blood and bone marrow containing allografts. Recommendations concerned donor screening, testing and re-testing of living donors after 6 months. The constant update for screening of donors for infectious diseases proved to be important in the subsequent years with two reports on transmission of hepatitis C virus by bone allografts (Eggen & Nordbø 1992; Conrad et al 1995). In the following years tissue banks developed better techniques for processing and preparation of bone allografts and more reliable blood tests came to market (Busch 1991, 1994; Alter et al. 1990). However, donor screening methods are constantly updated and revised with the introduction of new infectious diseases, like SARS (Lam et al. 2004). The safety of allograft bone transplants can never be taken for granted, but recent safety records for bone allografts are excellent. As more complex orthopaedic surgical procedures are performed nowadays the need for

### **3. Indications for the use of bone allograft**

(safe) bone allografts has increased (Nielsen et al. 2001).

Massive bone defects can arise from trauma, infection, osteolysis after arthroplasty or resection of bone tumours and are a challenging problem in orthopaedic practice. These bone defects can be filled with either autograft or allograft bone transplants. Ideally, autograft is preferred because of its osteoconductive and osteoinductive activity. However, autografts are available in limited number and size and therefore not sufficiently available. In addition harvesting is associated with extended surgical time and involves donor site morbidity (Aro&Aho 1993; Summers and Eisenstein 1989). Therefore allografts supplied by a bone bank are commonly used instead. Allogenic bone exclusively has osteoconductive activity; it serves as an acellular mineralized frame against which newly formed bone gets deposited (Elves and Pratt 1975; Urist 1953). Indications for the use of allografts are wide

Quality Control in Hospital Bone Banking 255

A bone bank procedure should be extensively described in a protocol concerning the five components: organization, donor selection, documentation, storage and processing, and implementation. The Head of Department (HOD). of the Department of Orthopaedics and

In an organization chart we describe the responsibilities of different stakeholders. One of the orthopaedic departmental members, preferably the HOD of Orthopaedic surgery, has general responsibility for the bone bank. The HOD should have appropriate technical support by a bone bank administrator, who is responsible for the daily management of the bank. This bone bank administrator can be a paramedic with appropriate training for the required tasks. The bone bank administrators' responsibilities include administration as well as storage and allocation of donor bone. Additionally, the administrator takes care of the maintenance and cleaning of the storage facilities (freezers, etc.), and verifies the registration forms of femoral heads meeting the requirements for storage in the bone bank. Furthermore the team consists of a theatre nurse, a medical microbiologist, an anatomic pathologist, a clinical chemical analyst, a haematological laboratory technician, and a trainer. The knowledge and skills concerning surgical techniques and clinical hygiene are guaranteed by the orthopaedic surgeon and theatre nurse. The bone bank administrator and the trainer are responsible for training of bone bank employees. Apart from an orientation module for new employees, the training program consists of regular refresher courses for all members of the

Preceding the hip replacement procedure, the attending orthopaedic surgeon requests the patient for his permission to store any removed tissue for donation. It concerns patients

prosthesis. The quantity of corticospongious bone removed during knee or shoulder arthroplasty is not sufficient for donor purposes; therefore patients undergoing such

The attending orthopaedic surgeon informs the patient both orally and written. In case the patient grants permission he or she signs the consent forms, and fills out a standard survey (see Table 1). The orthopedic surgeon now decides whether the patient is suitable for being

All criteria must be met; if not, exclusion necessarily follows. The orthopedic surgeon examines the patient thoroughly: blood samples are collected to determine blood type,

During surgery, bacterial culture swab samples from the hip capsule are collected and a biopsy of 1 cm3 corticospongious bone is sent for histopathological analysis. Serological screening for infectious diseases is performed six months after surgery. Once all requirements are met (Tables 1, 2, 3, 4, 5), a femoral head can be released for donation:

whose femoral head grafts will be retrieved in order to be replaced by a total hip

procedures cannot be taken into consideration for donation of bone tissue.

a donor; he uses general and specific exclusion criteria (see Tables 2, 3).

Rhesus-factor and Erythrocyte Sedimentation Rate (ESR). (Tables 4, 5).

• completed survey; all questions should yield a negative answer

• no abnormal bacteriological values in derived tissue

**4. Hospital bone bank protocol**

**4.1 Organization**

**4.2 Donor selection**

• approval donor

• signed consent forms of donor

• preoperative ESR rates within criteria

the bone bank administrator compose this protocol.

staff, in order to keep the knowledge of the procedures updated.

and include treatment of bone defects as a result from trauma, tumour surgery and infection (Friedlaender 1987; Mankin et al. 1976, 1996; Finkemeier 2002; Jupiter et al. 1987). In addition it is used in spinal fusion (Raizman et al. 2009; Takaso et al. 2011). However the most applied indication is the use of morselized allograft femoral heads as impaction grafting in revision or primary total joint arthroplasty. Aseptic loosening of the acetabular or femoral component of a total hip prosthesis is becoming an increasingly significant problem in orthopaedic surgery (Slooff et al. 1996). The migration of implants during loosening and procedures to remove the prosthesis and cement during revision induce significant bone destruction, resulting in enlargement of the acetabulum and widening of the femoral medullary cavity. Because of the magnitude of the loss of bone, allograft bone is often needed, at revision, to provide stability for the new socket or stem. Using a technique for revision of the acetabular component with impacted morselized cancellous bone grafting and a cemented or uncemented acetabular component, excellent long term results were described, with a survival of the revised hip prostheses of 87% at 20 years. (Schreurs et al. 2009; Garcia-Cimbrelo 2010; Paxton 2011). In different subgroups of patients this technique was applied with good results: patients under the age of 50, with rheumatoid arthritis, with acetabular fracture and dysplastic hips. In addition femoral revision with use of an impaction bone-grafting technique and a cemented polished stem resulted in an excellent prosthetic survival rate at eight to thirteen years postoperatively. However several studies report on high complication rates on this procedure, mostly femoral fractures and authors refined their indications (Leopold et al. 1999; Meding et al. 1997; Toms et al. 2004; Sierra et al. 2008). Because of the use of femoral head allograft bone in revision hip surgery, which is a common orthopaedic operation, the need for these allografts is increased. The retrieval of these femoral heads in daily orthopaedic practice is simple: the procedure for implantation of a total hip always requires removal of the femoral head. However the bone banking procedure for safe donor selection, retrieval, documentation and storage is a complex process. Therefore in many countries central bone banks were founded, containing boneand tendon allografts of various sizes and origin, obtained from living and deceased donor patients. These central bone banks also provide femoral head allografts to other hospitals, which can be ordered at a cost price in advance of a planned operation.

Hospitals that perform a high quantity of orthopaedic procedures that require the use of bone allografts (total hip revisions, spine surgery and tumour surgery) might consider foundation of a hospital owned bone bank. In the Netherlands these hospital bone banks contain only allografts of femoral heads of living donors removed at the time of hip replacement surgery. Main advantages for the hospital and orthopaedic surgeons consist of easy accessibility to - and availability of allografts. In addition there might be a financial advantage, depending on the quantity of allografts needed per year. However the breakeven point is not clear and dependant on local costs which differ between hospitals, countries and bone banking protocols. In the next paragraph a bone banking protocol is described which meets the requirements of Dutch national law and European guidelines 2004/23/EC and 2006/86/EC and is based on several guidelines: The American Association of Tissue Banks (AATB 1993), the criteria of the Council for Blood Transfusion of the Netherlands Red Cross (Richtlijn Bloedtransfusie 2004), the central bone bank in the Netherlands NBF-BIS (NBF-BIS Foundation 2010) and the guidelines of the European Association of Musculoskeletal Transplantation (EAMST), which has been discontinued because of diverging European legislation.

#### **4. Hospital bone bank protocol**

A bone bank procedure should be extensively described in a protocol concerning the five components: organization, donor selection, documentation, storage and processing, and implementation. The Head of Department (HOD). of the Department of Orthopaedics and the bone bank administrator compose this protocol.

#### **4.1 Organization**

254 Wide Spectra of Quality Control

and include treatment of bone defects as a result from trauma, tumour surgery and infection (Friedlaender 1987; Mankin et al. 1976, 1996; Finkemeier 2002; Jupiter et al. 1987). In addition it is used in spinal fusion (Raizman et al. 2009; Takaso et al. 2011). However the most applied indication is the use of morselized allograft femoral heads as impaction grafting in revision or primary total joint arthroplasty. Aseptic loosening of the acetabular or femoral component of a total hip prosthesis is becoming an increasingly significant problem in orthopaedic surgery (Slooff et al. 1996). The migration of implants during loosening and procedures to remove the prosthesis and cement during revision induce significant bone destruction, resulting in enlargement of the acetabulum and widening of the femoral medullary cavity. Because of the magnitude of the loss of bone, allograft bone is often needed, at revision, to provide stability for the new socket or stem. Using a technique for revision of the acetabular component with impacted morselized cancellous bone grafting and a cemented or uncemented acetabular component, excellent long term results were described, with a survival of the revised hip prostheses of 87% at 20 years. (Schreurs et al. 2009; Garcia-Cimbrelo 2010; Paxton 2011). In different subgroups of patients this technique was applied with good results: patients under the age of 50, with rheumatoid arthritis, with acetabular fracture and dysplastic hips. In addition femoral revision with use of an impaction bone-grafting technique and a cemented polished stem resulted in an excellent prosthetic survival rate at eight to thirteen years postoperatively. However several studies report on high complication rates on this procedure, mostly femoral fractures and authors refined their indications (Leopold et al. 1999; Meding et al. 1997; Toms et al. 2004; Sierra et al. 2008). Because of the use of femoral head allograft bone in revision hip surgery, which is a common orthopaedic operation, the need for these allografts is increased. The retrieval of these femoral heads in daily orthopaedic practice is simple: the procedure for implantation of a total hip always requires removal of the femoral head. However the bone banking procedure for safe donor selection, retrieval, documentation and storage is a complex process. Therefore in many countries central bone banks were founded, containing boneand tendon allografts of various sizes and origin, obtained from living and deceased donor patients. These central bone banks also provide femoral head allografts to other hospitals,

which can be ordered at a cost price in advance of a planned operation.

because of diverging European legislation.

Hospitals that perform a high quantity of orthopaedic procedures that require the use of bone allografts (total hip revisions, spine surgery and tumour surgery) might consider foundation of a hospital owned bone bank. In the Netherlands these hospital bone banks contain only allografts of femoral heads of living donors removed at the time of hip replacement surgery. Main advantages for the hospital and orthopaedic surgeons consist of easy accessibility to - and availability of allografts. In addition there might be a financial advantage, depending on the quantity of allografts needed per year. However the breakeven point is not clear and dependant on local costs which differ between hospitals, countries and bone banking protocols. In the next paragraph a bone banking protocol is described which meets the requirements of Dutch national law and European guidelines 2004/23/EC and 2006/86/EC and is based on several guidelines: The American Association of Tissue Banks (AATB 1993), the criteria of the Council for Blood Transfusion of the Netherlands Red Cross (Richtlijn Bloedtransfusie 2004), the central bone bank in the Netherlands NBF-BIS (NBF-BIS Foundation 2010) and the guidelines of the European Association of Musculoskeletal Transplantation (EAMST), which has been discontinued In an organization chart we describe the responsibilities of different stakeholders. One of the orthopaedic departmental members, preferably the HOD of Orthopaedic surgery, has general responsibility for the bone bank. The HOD should have appropriate technical support by a bone bank administrator, who is responsible for the daily management of the bank. This bone bank administrator can be a paramedic with appropriate training for the required tasks. The bone bank administrators' responsibilities include administration as well as storage and allocation of donor bone. Additionally, the administrator takes care of the maintenance and cleaning of the storage facilities (freezers, etc.), and verifies the registration forms of femoral heads meeting the requirements for storage in the bone bank. Furthermore the team consists of a theatre nurse, a medical microbiologist, an anatomic pathologist, a clinical chemical analyst, a haematological laboratory technician, and a trainer. The knowledge and skills concerning surgical techniques and clinical hygiene are guaranteed by the orthopaedic surgeon and theatre nurse. The bone bank administrator and the trainer are responsible for training of bone bank employees. Apart from an orientation module for new employees, the training program consists of regular refresher courses for all members of the staff, in order to keep the knowledge of the procedures updated.

#### **4.2 Donor selection**

Preceding the hip replacement procedure, the attending orthopaedic surgeon requests the patient for his permission to store any removed tissue for donation. It concerns patients whose femoral head grafts will be retrieved in order to be replaced by a total hip

prosthesis. The quantity of corticospongious bone removed during knee or shoulder arthroplasty is not sufficient for donor purposes; therefore patients undergoing such procedures cannot be taken into consideration for donation of bone tissue.

The attending orthopaedic surgeon informs the patient both orally and written. In case the patient grants permission he or she signs the consent forms, and fills out a standard survey (see Table 1). The orthopedic surgeon now decides whether the patient is suitable for being a donor; he uses general and specific exclusion criteria (see Tables 2, 3).

All criteria must be met; if not, exclusion necessarily follows. The orthopedic surgeon examines the patient thoroughly: blood samples are collected to determine blood type, Rhesus-factor and Erythrocyte Sedimentation Rate (ESR). (Tables 4, 5).

During surgery, bacterial culture swab samples from the hip capsule are collected and a biopsy of 1 cm3 corticospongious bone is sent for histopathological analysis. Serological screening for infectious diseases is performed six months after surgery. Once all requirements are met (Tables 1, 2, 3, 4, 5), a femoral head can be released for donation:

Quality Control in Hospital Bone Banking 257

Retrieval of the femoral head is performed at the time of routine orthopaedic total hip replacement under aseptic conditions. The removed femoral head is inspected and capsule and synovial tissue are cultured on aerobic and anaerobic bacteria. In order to exclude malignancies, auto-immune processes, or infections, a biopsy of 1 cm3 corticospongious bone and capsule is collected for histopathological examination. After determining its size,

continuous temperature registration device installed. Should the temperature fall outside the acceptable range of -90 and -70°C, an alarm system gives off a warning signal to the Technical Service, guaranteeing a 24- hours security against temperature-induced damage to the tissue. A nitrogen tank is fitted onto the freezer, as a backup cooling mechanism in case of mechanical breakdown of the freezer. In deep frozen condition, the allogenic bone tissue can be preserved for a maximum of 5 years. The temperature data is stored and

Anamnesis of previous infection with hepatitis B or C, AIDS or AIDS related complex, or tested

Recent (<4 weeks). vaccination with live vaccine (measles, yellow fever, mumps, polio, oral typhoid,

Language barrier or when patient does not understand the information for any reason (e.g.

If during surgery an orthopaedic surgeon decides to use a femoral head as an allograft, a femoral head from the freezer together with its documents are handed over to the orthopedic surgeon and surgery team. The orthopedic surgeon and theatre nurse verify the file and expiration date of the femoral head. The femoral head is thawed in physiological

saline; after being defrosted the theatre nurse takes a bacterial culture swab.

the femoral head is wrapped in three layers of sterile packing material, labeled and stored in the freezer within 30 min. The freezer has a temperature of -80°C, and has a

managed by the bone bank administrator for a period of at least 5 years.

Active infection of transplantation tissue (especially coxitis/osteomyelitis).

**4.4 Storage and processing**

No permission from patient Under aged donor (<18 years).

positive for HIV

Rheumatoid arthritis

Metabolic disorders

psychiatric patients).

Malignancies

Dementia

rubella).

Active or recent systemic infection/sepsis

Active ''slow-virus'' infection or anamnesis in the past

Diffuse connective tissue disorders/autoimmune diseases

Existing insulin dependent diabetes mellitus

Chronic medication (especially corticosteroids).

Donor location has been exposed to radiation

Previously infected with tuberculosis

Active or past syphilis infection

Treatment with growth hormones

Recent exposure to toxic substances

Table 2. General exclusion criteria

**4.5 Allocation and implementation**

Chronic neurological disorders

#### **4.3 Documentation**

Accurate documentation and coding are of the utmost importance for a well functioning bone bank. A unique registration code is allocated to each femoral head. Only the bone bank administrator is able to trace the donor based on this code. Of every registered femoral head, a file, containing the consent forms and results of ESR, bacteriological and histopathological examination, is kept updated. Other relevant data, such as the size of the femoral head and the allocation date are also documented and stored in this file. When the file is completed (which takes at least 6 months due to the serological examination), and no abnormalities are recorded, both bone bank administrator and the responsible orthopedic surgeon sign the forms. The femoral head is now available for transplantation. In case a file cannot be completed in full, or any abnormal values are recorded, the femoral head will be destroyed according to hospitals' protocol.

In the past 3 months, did you suffer any infection? If so, what infection? In the past 3 months, did you have any vaccination or inoculation, or have you been injected with narcotic drugs? In the past 6 months, did you have a malaria attack or did you use anti-malarial medication?  Have you ever been diagnosed with jaundice or liver illness? In the past 6 months, have you been in contact with patients diagnosed with jaundice/hepatitis? In the past 6 months, have you been in contact with patients diagnosed with AIDS, or individuals at risk to AIDS? If yes, how and when? Have you ever been tested for HIV/AIDS?  (Males only). Have you emigrated after 1977? If so, to what country? Are you diagnosed with hemophilia? If yes, are you using anticoagulants? Are you a sexual partner of an individual for which any of the abovementioned questions can be answered with 'yes'? Have you been actively involved in prostitution after 1977, or have you been a sexual partner of a person involved in prostitution in the past 6 months? Have you ever been diagnosed with a hematological disease or any malignant disorder? Have you ever been treated for diabetes mellitus? Have you ever been treated for chronic brain- or neurological diseases? Have you ever received radiation therapy? Have you ever been diagnosed with rheumatoid arthritis? Have you ever been diagnosed with tuberculosis? Have you ever been diagnosed with any disease, other than the abovementioned? Have you ever received hormonal treatment? Do you use any prescribed medication? Have you ever used any narcotic drugs? Have you recently been exposed to hazardous or toxic materials? If yes, please specify. What is your alcohol consumption per week? Have you recently been in surgery? If so, when? Did you receive blood from a blood transfusion? In the past 14 days, have you been traveling through or staying in a region exposed to a SARS epidemic, or have you been in contact with patients infected with SARS? In the past 6 months, have you tattooed yourself or did you get a piercing? Table 1. Questionnaire patient for orthopaedic bone donation

#### **4.4 Storage and processing**

256 Wide Spectra of Quality Control

Accurate documentation and coding are of the utmost importance for a well functioning bone bank. A unique registration code is allocated to each femoral head. Only the bone bank administrator is able to trace the donor based on this code. Of every registered femoral head, a file, containing the consent forms and results of ESR, bacteriological and histopathological examination, is kept updated. Other relevant data, such as the size of the femoral head and the allocation date are also documented and stored in this file. When the file is completed (which takes at least 6 months due to the serological examination), and no abnormalities are recorded, both bone bank administrator and the responsible orthopedic surgeon sign the forms. The femoral head is now available for transplantation. In case a file cannot be completed in full, or any abnormal values are recorded, the femoral head will be

In the past 3 months, did you have any vaccination or inoculation, or have you been injected with

In the past 6 months, have you been in contact with patients diagnosed with jaundice/hepatitis?

Have you ever been diagnosed with any disease, other than the abovementioned?

Have you recently been exposed to hazardous or toxic materials? If yes, please specify.

Have you recently been in surgery? If so, when? Did you receive blood from a blood transfusion? In the past 14 days, have you been traveling through or staying in a region exposed to a SARS

In the past 6 months, did you have a malaria attack or did you use anti-malarial medication?

• no abnormal histopathological structures in derived tissue • no abnormal serological values six months after surgery

In the past 3 months, did you suffer any infection? If so, what infection?

 Have you ever been diagnosed with jaundice or liver illness?

 (Males only). Have you emigrated after 1977? If so, to what country?

person involved in prostitution in the past 6 months?

Have you ever been diagnosed with rheumatoid arthritis? Have you ever been diagnosed with tuberculosis?

Are you diagnosed with hemophilia? If yes, are you using anticoagulants?

Have you ever been treated for chronic brain- or neurological diseases?

epidemic, or have you been in contact with patients infected with SARS? In the past 6 months, have you tattooed yourself or did you get a piercing?

Table 1. Questionnaire patient for orthopaedic bone donation

**4.3 Documentation**

narcotic drugs?

answered with 'yes'?

destroyed according to hospitals' protocol.

 risk to AIDS? If yes, how and when? Have you ever been tested for HIV/AIDS?

Have you ever been treated for diabetes mellitus?

Have you ever received radiation therapy?

Have you ever received hormonal treatment? Do you use any prescribed medication? Have you ever used any narcotic drugs?

What is your alcohol consumption per week?

Retrieval of the femoral head is performed at the time of routine orthopaedic total hip replacement under aseptic conditions. The removed femoral head is inspected and capsule and synovial tissue are cultured on aerobic and anaerobic bacteria. In order to exclude malignancies, auto-immune processes, or infections, a biopsy of 1 cm3 corticospongious bone and capsule is collected for histopathological examination. After determining its size, the femoral head is wrapped in three layers of sterile packing material, labeled and stored in the freezer within 30 min. The freezer has a temperature of -80°C, and has a continuous temperature registration device installed. Should the temperature fall outside the acceptable range of -90 and -70°C, an alarm system gives off a warning signal to the Technical Service, guaranteeing a 24- hours security against temperature-induced damage to the tissue. A nitrogen tank is fitted onto the freezer, as a backup cooling mechanism in case of mechanical breakdown of the freezer. In deep frozen condition, the allogenic bone tissue can be preserved for a maximum of 5 years. The temperature data is stored and managed by the bone bank administrator for a period of at least 5 years.

No permission from patient Under aged donor (<18 years). Active or recent systemic infection/sepsis Active infection of transplantation tissue (especially coxitis/osteomyelitis). Previously infected with tuberculosis Active ''slow-virus'' infection or anamnesis in the past Anamnesis of previous infection with hepatitis B or C, AIDS or AIDS related complex, or tested positive for HIV Active or past syphilis infection Recent (<4 weeks). vaccination with live vaccine (measles, yellow fever, mumps, polio, oral typhoid, rubella). Rheumatoid arthritis Diffuse connective tissue disorders/autoimmune diseases Metabolic disorders Existing insulin dependent diabetes mellitus Treatment with growth hormones Chronic medication (especially corticosteroids). Recent exposure to toxic substances Malignancies Donor location has been exposed to radiation Chronic neurological disorders Dementia Language barrier or when patient does not understand the information for any reason (e.g. psychiatric patients).

Table 2. General exclusion criteria

#### **4.5 Allocation and implementation**

If during surgery an orthopaedic surgeon decides to use a femoral head as an allograft, a femoral head from the freezer together with its documents are handed over to the orthopedic surgeon and surgery team. The orthopedic surgeon and theatre nurse verify the file and expiration date of the femoral head. The femoral head is thawed in physiological saline; after being defrosted the theatre nurse takes a bacterial culture swab.

Quality Control in Hospital Bone Banking 259

The bone bank protocol contains several procedures to secure the retrieval, storage and implementation of predictable, safe femoral head allografts. To prevent transmission of infectious diseases and malignancies not only thorough donor screening, but also histopathological screening, culture swabs and serological screening are performed as a tool for quality control. In literature these complementary screening methods are discussed.

There is no uniform practice concerning histopathological evaluation of femoral heads retrieved from living donors. Some hospitals routinely examine all removed tissue specimens histopathological, suitable for bone banking or not. At some only gross examination is performed and at some neither evaluation is done. The utility and costperformance ratio of routine histopathological evaluation is discussed in literature. Several studies doubt the cost effectiveness and necessity of routine histopathological examination (Campbell et al. 1997; Kocher et al. 2000; Lawrence et al. 1999; Meding et al. 2000; Raab et al. 1998). Campbell et al. performed a retrospective review of 715 pathologic findings of specimens obtained at total joint arthroplasties (283 hips and 432 knees). Of 715 specimens 3 were suggestive for low grade lympho-proliferative disorder and 3 for rheumatoid arthritis. In none of these cases this resulted in alteration in patient care. Follow up by telephone at 35 months revealed no signs of systemic disease. Therefore they conclude that routine histopathologic evaluation is not cost effective (Campbell et al. 1997). Kocher et al. performed 1234 arthroplasties (471 hips 763 knees) and found discrepant diagnoses in 28 patients, including (pseudo)gout, pigmented villonodular synovitis, osteonecrosis, granulomatous disease, hemochromatosis and rheumatoid arthritis. In two patients they found hypercellular marrow suggestive for lympho- proliferative disease. They performed no formal hematological work-up. They conclude that because of low prevalence of findings that altered patient management, routine pathological examination has limited cost effectiveness (Kocher et al. 1998). Meding et al. drawed similar conclusions. They found 27 discrepancies in pathology in 951 cases, however no neoplasia was noted. Lawrence et al. found malignancies in 11 of 1388 patients, including three cases of lympho- proliferative disease. They stated that the surgeon was able to identify malignancy with a sensitivity of 100 % and a specificity of 99.9%. They conclude that the surgeon preoperatively can exclude the possibility of malignancy and should reserve histopathological analysis for cases where the diagnosis is suspect or in case of unsuspected intra-operative findings (Lawrence et al. 1999). Raab et al. found two malignancies in a group of 168 patients, one of which had findings suggestive for lympho-proliferative disorder, however patient management did not alter. One patient showed osteomyelitis and this had clinical significance for patient treatment. In retrospect this turned out to be a misdiagnosis (Raab et al. 1998). The authors of these studies state that omission of routine pathology would bring about substantial decreases in charges. Campbell et al. estimated that the potential annual cost savings would be between \$43 million to \$61 million if all hip and knee arthroplasties in the USA were considered. Kocher et al. reported on \$122,000 in total costs for them to discover only one unexpected case of granulomatous inflammation. Total reimbursements were only slightly more than \$106,000, leaving them with a \$16,000 loss for the hospital. Clearly, desisting with routine pathology for presumed osteoarthritis in elective total joint cases could substantially decrease expenditures. The earlier described studies make compelling arguments for omission

**5. Quality control in hospital bone banking**

**5.1 Histopathological screening**

The hospital or care institution warrants fulfillment of the traceability requirements, which implies storing the file of the femoral head and records of the receiving patient for 30 years post implantation.

A clinically proven HIV infection Men having homosexual intercourse after 1977 Intravenous medication/narcotics use, currently or in the past Immigrants (after 1977). from countries of which it is known that heterosexual intercourse is an important factor for HIV transmission Hemophilic patients administered with clotting factors concentrate Sexual partners from abovementioned individuals Men and women active in prostitution since 1977, and individuals being their partner in the past 6 months Individuals who recently (past 6 months). placed a tattoo or piercing Individuals who have had a blood transfusion before 1980 Individuals who have had a blood transfusion outside Europe or North America Individuals who stayed in a SARS epidemic area or individuals who had face-to-face contact with a SARS patient

Table 3.  Hematological examination before surgery

Viral (hepatitis). Hepatitis B antigen Hepatitis B antibody Hepatitis C antibody Viral nucleic acid Viral (additional). HIV HTLV Bacterial syphilis TPHA (Treponema Pallidum, Lues).

Table 5. Serological examination 6 months postoperative

The hospital or care institution warrants fulfillment of the traceability requirements, which implies storing the file of the femoral head and records of the receiving patient for 30 years

Immigrants (after 1977). from countries of which it is known that heterosexual intercourse is an

Men and women active in prostitution since 1977, and individuals being their partner in the past 6

Individuals who stayed in a SARS epidemic area or individuals who had face-to-face contact with a

post implantation.

months

SARS patient

A clinically proven HIV infection

important factor for HIV transmission

Table 3. Specific exclusion criteria

Blood type and Rhesus factor

Normal values Male

Female

 <50 years: 0–15 >50 years: 0–20

 <50 years: 0–20 >50 years: 0–30

Viral (hepatitis). Hepatitis B antigen Hepatitis B antibody Hepatitis C antibody Viral nucleic acid Viral (additional).

TPHA (Treponema Pallidum, Lues).

 HIV HTLV Bacterial syphilis

Men having homosexual intercourse after 1977

Sexual partners from abovementioned individuals

Intravenous medication/narcotics use, currently or in the past

Hemophilic patients administered with clotting factors concentrate

Individuals who recently (past 6 months). placed a tattoo or piercing

Individuals who have had a blood transfusion outside Europe or North America

Individuals who have had a blood transfusion before 1980

Erythrocyte sedimentation rate (ESR), age and sex dependant

Table 4. Hematological examination before surgery

Table 5. Serological examination 6 months postoperative

#### **5. Quality control in hospital bone banking**

The bone bank protocol contains several procedures to secure the retrieval, storage and implementation of predictable, safe femoral head allografts. To prevent transmission of infectious diseases and malignancies not only thorough donor screening, but also histopathological screening, culture swabs and serological screening are performed as a tool for quality control. In literature these complementary screening methods are discussed.

#### **5.1 Histopathological screening**

There is no uniform practice concerning histopathological evaluation of femoral heads retrieved from living donors. Some hospitals routinely examine all removed tissue specimens histopathological, suitable for bone banking or not. At some only gross examination is performed and at some neither evaluation is done. The utility and costperformance ratio of routine histopathological evaluation is discussed in literature. Several studies doubt the cost effectiveness and necessity of routine histopathological examination (Campbell et al. 1997; Kocher et al. 2000; Lawrence et al. 1999; Meding et al. 2000; Raab et al. 1998). Campbell et al. performed a retrospective review of 715 pathologic findings of specimens obtained at total joint arthroplasties (283 hips and 432 knees). Of 715 specimens 3 were suggestive for low grade lympho-proliferative disorder and 3 for rheumatoid arthritis. In none of these cases this resulted in alteration in patient care. Follow up by telephone at 35 months revealed no signs of systemic disease. Therefore they conclude that routine histopathologic evaluation is not cost effective (Campbell et al. 1997). Kocher et al. performed 1234 arthroplasties (471 hips 763 knees) and found discrepant diagnoses in 28 patients, including (pseudo)gout, pigmented villonodular synovitis, osteonecrosis, granulomatous disease, hemochromatosis and rheumatoid arthritis. In two patients they found hypercellular marrow suggestive for lympho- proliferative disease. They performed no formal hematological work-up. They conclude that because of low prevalence of findings that altered patient management, routine pathological examination has limited cost effectiveness (Kocher et al. 1998). Meding et al. drawed similar conclusions. They found 27 discrepancies in pathology in 951 cases, however no neoplasia was noted. Lawrence et al. found malignancies in 11 of 1388 patients, including three cases of lympho- proliferative disease. They stated that the surgeon was able to identify malignancy with a sensitivity of 100 % and a specificity of 99.9%. They conclude that the surgeon preoperatively can exclude the possibility of malignancy and should reserve histopathological analysis for cases where the diagnosis is suspect or in case of unsuspected intra-operative findings (Lawrence et al. 1999). Raab et al. found two malignancies in a group of 168 patients, one of which had findings suggestive for lympho-proliferative disorder, however patient management did not alter. One patient showed osteomyelitis and this had clinical significance for patient treatment. In retrospect this turned out to be a misdiagnosis (Raab et al. 1998). The authors of these studies state that omission of routine pathology would bring about substantial decreases in charges. Campbell et al. estimated that the potential annual cost savings would be between \$43 million to \$61 million if all hip and knee arthroplasties in the USA were considered. Kocher et al. reported on \$122,000 in total costs for them to discover only one unexpected case of granulomatous inflammation. Total reimbursements were only slightly more than \$106,000, leaving them with a \$16,000 loss for the hospital. Clearly, desisting with routine pathology for presumed osteoarthritis in elective total joint cases could substantially decrease expenditures. The earlier described studies make compelling arguments for omission

Quality Control in Hospital Bone Banking 261

and perform routine histopathological screening as part of the bone banking protocol

In order to prevent transplantation of infected bone allografts we routinely perform culture swabs of the femoral head and synovium. If these culture swabs are positive for bacterial contamination the femoral head is discarded. Because femoral heads are readily available, any suspicion of contamination is respected, regardless of the source of the organism. At the time of implementation another two culture swabs from the thawed femoral head are performed. A culture of a specimen at the time of use of the femoral head serves two purposes: it is a quality-control check on the banking procedure, without the risk of additional contamination by separate culturing and handling, and it also allows the surgeon to administer an appropriate antibiotic should the culture be positive, especially if an infection occurs postoperatively (Tomford et al. 1986). However literature suggests that

Veen et al (1994) describe analysis of 75 fibular specimens obtained from cadaver donors under sterile conditions. All specimens were culture swabbed as routinely performed for retrieved allografts. Of these allografts 92 % were contaminated when cultured entirely but swab cultures were positive in only 45% After swabbing, all specimens were placed in BHIculture medium. Three different protocols were subsequently followed: 1) culture of the entire bone specimen in BHI-culture medium, 2) culture of the swab incubated on blood agar and chocolate plates, and 3) culture of the swab in BHI-culture medium. A control group included 20 sterilized bone specimens that were cultured entirely according to Protocol 1. The negative predictive value and sensitivity and were found to be 9% and 10% in Protocol 2 and 13% in Protocol 3. Therefore they conclude that swab cultures are inadequate to detect bacterial contamination of bone allografts in all cases. However, because of an acceptable infection rate after transplantation of the allografts that does not exceed those reported in other similar series, there is suggestion of an acceptable bioburden. Vehmeyer et al (2002) analyzed the bacterial contamination of 106 allografts of femoral heads obtained from living donors. From 15 initially swab positive grafts only five grafts were contaminated when cultured entirely. From 10 of 91 initially swab culture negative allografts microorganisms could be isolated when cultered in their entirety. They conclude that the routine swab culture technique seems to be less suitable for assessing the bacterial load of femoral heads obtained from living donors. Therefore they advise to routinely

routine culture swabs are not always able to detect bacterial contamination.

perform antibacterial processing before releasing an allograft for transplantation.

allografts obtained from post-mortem donors (Deijkers et al. 1997).

clinical implications for the donor.

**5.3 Immunogenic screening**

Antibiotic rinsing of the allograft seems not to be an effective decontamination method in

James et al. (2002; 2004) determined whether the swab culture results had any clinical implication on wound problems or infections in the donor. In performed studies the rate of contamination was 9%, which is consistent with other studies. There was no difference in the complication rate of patients with a positive culture swab compared to those with a negative culture swab and therefore they conclude that positive culture swabs have no

A question of interest to all bone banks was raised by a case report of a young Rhesusnegative female patient in whom antibodies to a Rhesus antigen developed after she

(Zwitser et al. 2009).

**5.2 Culture swabs**

of routine pathologic examination after elective joint arthroplasty for presumed osteoarthritis. The cost savings they estimate are enormous and patient management did not change because of an unexpected histologic diagnosis.

On the other hand some studies find higher percentages of unexpected pathological diagnoses and therefore advise routine histopathological examination (Billings et al. 2000; Clark et al. 2000; DiCarlo et al. 1994; Lauder et al. 2004; Palmer et al. 1999; Sugihara et al. 1999; Zwitser et al. 2009). DiCarlo et al. documented a 5.4% disagreement between the clinical and pathologic diagnoses of 1794 femoral heads from total joint arthroplasties , with a large-cell lymphoma, myeloma, sarcoma, ochronosis, Gaucher's disease, Paget's disease, and enchondroma. The authors recommended routine pathology for elective arthroplasty to both verify the diagnosis and to serve as a measure of quality control. Billings et al. described the unexpected finding of an occult primary bone sarcoma in two patients with otherwise benign clinical findings and therefore underscore the necessity for routine pathological examination of femoral head specimens from patients who are at risk for the development of a secondary malignant tumor (Billings et al. 2000).

Lauder et al. describe a patient with histopathological findings indicative for a low grade B cell lymphoma who developed systemic disease after 8 months. They underscore the fact that without routine pathologic examination, neoplasms could still be missed, even in patients who lack risk factors for malignancy and despite of a thorough preoperative evaluation. Furthermore they discuss the fact that studies in which low grade malignancies were found, suggested that their patients had been free of signs or symptoms for underlying disease, but did not perform a formal hematological evaluation for malignancy (Kocher et al. 2000; Campbell et al. 1997). They ask what the cost-benefit is when one is able to diagnose an unsuspected occult malignancy (Lauder et al. 2004). Clark et al. conclude that the routine histological evaluation of tissue excised from patients with an uncomplicated case of osteoarthritis may not be necessary at all hospitals, but when a patient has suspection of another disorder then osteoarthritis and when gross examination suggests an unexpected finding, or when the results of such analysis are used for ongoing quality-assurance studies, histological examination is warranted. Other studies performed analysis of histopathological screening of femoral heads for bone banking screening purposes.

 Palmer et al. analysed the histological findings in 1146 osteoarthritic femoral heads which would have been considered suitable for bone bank donation to determine presence of pathological lesions and found that 91 femoral heads (8%) showed evidence of disease. The most common benign conditions were chondrocalcinosis (63), avascular necrosis (13), osteomas (6), metabolic bone disease (2) and rheumatoid arthritis (4). Three cases of malignant tumour were described (one case of low-grade chondrosarcoma and two of welldifferentiated lymphocytic lymphoma). They conclude that occult pathological conditions are common and recommend that histopathological screening should be included as part of the screening protocol for bone bank collection (Palmer et al. 1999). Sugihara et al. describe similar histological findings in routine bone bank screening of 137 femoral heads and found abnormal histopathological findings in five femoral heads: three were highly suspicious of low-grade B-cell lymphoma, one of monoclonal plasmacytosis and the other of non-specific inflammation of bone marrow (Sugihara et al.,1999). In routine histopathological screening the subsequent years this group found variable numbers of low-grade B-cell lymphoma, even in a group of femoral heads that were eligible for bone transplantation. In a long term follow up of these patients, with serendipitously found low grade B cell lymphoma on routine histologic examination, two developed systemic disease. Therefore we recommend and perform routine histopathological screening as part of the bone banking protocol (Zwitser et al. 2009).

#### **5.2 Culture swabs**

260 Wide Spectra of Quality Control

of routine pathologic examination after elective joint arthroplasty for presumed osteoarthritis. The cost savings they estimate are enormous and patient management did not

On the other hand some studies find higher percentages of unexpected pathological diagnoses and therefore advise routine histopathological examination (Billings et al. 2000; Clark et al. 2000; DiCarlo et al. 1994; Lauder et al. 2004; Palmer et al. 1999; Sugihara et al. 1999; Zwitser et al. 2009). DiCarlo et al. documented a 5.4% disagreement between the clinical and pathologic diagnoses of 1794 femoral heads from total joint arthroplasties , with a large-cell lymphoma, myeloma, sarcoma, ochronosis, Gaucher's disease, Paget's disease, and enchondroma. The authors recommended routine pathology for elective arthroplasty to both verify the diagnosis and to serve as a measure of quality control. Billings et al. described the unexpected finding of an occult primary bone sarcoma in two patients with otherwise benign clinical findings and therefore underscore the necessity for routine pathological examination of femoral head specimens from patients who are at risk for the

Lauder et al. describe a patient with histopathological findings indicative for a low grade B cell lymphoma who developed systemic disease after 8 months. They underscore the fact that without routine pathologic examination, neoplasms could still be missed, even in patients who lack risk factors for malignancy and despite of a thorough preoperative evaluation. Furthermore they discuss the fact that studies in which low grade malignancies were found, suggested that their patients had been free of signs or symptoms for underlying disease, but did not perform a formal hematological evaluation for malignancy (Kocher et al. 2000; Campbell et al. 1997). They ask what the cost-benefit is when one is able to diagnose an unsuspected occult malignancy (Lauder et al. 2004). Clark et al. conclude that the routine histological evaluation of tissue excised from patients with an uncomplicated case of osteoarthritis may not be necessary at all hospitals, but when a patient has suspection of another disorder then osteoarthritis and when gross examination suggests an unexpected finding, or when the results of such analysis are used for ongoing quality-assurance studies, histological examination is warranted. Other studies performed analysis of histopathological

 Palmer et al. analysed the histological findings in 1146 osteoarthritic femoral heads which would have been considered suitable for bone bank donation to determine presence of pathological lesions and found that 91 femoral heads (8%) showed evidence of disease. The most common benign conditions were chondrocalcinosis (63), avascular necrosis (13), osteomas (6), metabolic bone disease (2) and rheumatoid arthritis (4). Three cases of malignant tumour were described (one case of low-grade chondrosarcoma and two of welldifferentiated lymphocytic lymphoma). They conclude that occult pathological conditions are common and recommend that histopathological screening should be included as part of the screening protocol for bone bank collection (Palmer et al. 1999). Sugihara et al. describe similar histological findings in routine bone bank screening of 137 femoral heads and found abnormal histopathological findings in five femoral heads: three were highly suspicious of low-grade B-cell lymphoma, one of monoclonal plasmacytosis and the other of non-specific inflammation of bone marrow (Sugihara et al.,1999). In routine histopathological screening the subsequent years this group found variable numbers of low-grade B-cell lymphoma, even in a group of femoral heads that were eligible for bone transplantation. In a long term follow up of these patients, with serendipitously found low grade B cell lymphoma on routine histologic examination, two developed systemic disease. Therefore we recommend

change because of an unexpected histologic diagnosis.

development of a secondary malignant tumor (Billings et al. 2000).

screening of femoral heads for bone banking screening purposes.

In order to prevent transplantation of infected bone allografts we routinely perform culture swabs of the femoral head and synovium. If these culture swabs are positive for bacterial contamination the femoral head is discarded. Because femoral heads are readily available, any suspicion of contamination is respected, regardless of the source of the organism. At the time of implementation another two culture swabs from the thawed femoral head are performed. A culture of a specimen at the time of use of the femoral head serves two purposes: it is a quality-control check on the banking procedure, without the risk of additional contamination by separate culturing and handling, and it also allows the surgeon to administer an appropriate antibiotic should the culture be positive, especially if an infection occurs postoperatively (Tomford et al. 1986). However literature suggests that routine culture swabs are not always able to detect bacterial contamination.

Veen et al (1994) describe analysis of 75 fibular specimens obtained from cadaver donors under sterile conditions. All specimens were culture swabbed as routinely performed for retrieved allografts. Of these allografts 92 % were contaminated when cultured entirely but swab cultures were positive in only 45% After swabbing, all specimens were placed in BHIculture medium. Three different protocols were subsequently followed: 1) culture of the entire bone specimen in BHI-culture medium, 2) culture of the swab incubated on blood agar and chocolate plates, and 3) culture of the swab in BHI-culture medium. A control group included 20 sterilized bone specimens that were cultured entirely according to Protocol 1. The negative predictive value and sensitivity and were found to be 9% and 10% in Protocol 2 and 13% in Protocol 3. Therefore they conclude that swab cultures are inadequate to detect bacterial contamination of bone allografts in all cases. However, because of an acceptable infection rate after transplantation of the allografts that does not exceed those reported in other similar series, there is suggestion of an acceptable bioburden. Vehmeyer et al (2002) analyzed the bacterial contamination of 106 allografts of femoral heads obtained from living donors. From 15 initially swab positive grafts only five grafts were contaminated when cultured entirely. From 10 of 91 initially swab culture negative allografts microorganisms could be isolated when cultered in their entirety. They conclude that the routine swab culture technique seems to be less suitable for assessing the bacterial load of femoral heads obtained from living donors. Therefore they advise to routinely perform antibacterial processing before releasing an allograft for transplantation.

Antibiotic rinsing of the allograft seems not to be an effective decontamination method in allografts obtained from post-mortem donors (Deijkers et al. 1997).

James et al. (2002; 2004) determined whether the swab culture results had any clinical implication on wound problems or infections in the donor. In performed studies the rate of contamination was 9%, which is consistent with other studies. There was no difference in the complication rate of patients with a positive culture swab compared to those with a negative culture swab and therefore they conclude that positive culture swabs have no clinical implications for the donor.

#### **5.3 Immunogenic screening**

A question of interest to all bone banks was raised by a case report of a young Rhesusnegative female patient in whom antibodies to a Rhesus antigen developed after she

Quality Control in Hospital Bone Banking 263

practice is extensively discussed on in literature. At this moment we have no financially viable bone bank. This is due to organisational and logistical problems, which have our

Alter HJ, Epstein JS, Swenson SG, VanRaden MJ, Ward JW, Kaslow RA, Menitove JE, Klein

American Association of Tissue Banks (AATB). (1993). In: *Standards for tissue banking.*

Billings S.D, Wurtz L.D, Tejada E & Henley J.D (2000). Occult sarcoma of the femoral head

Busch MP, Eble BE, Khayam-Bashi H, Heilbron D, Murphy EL, Kwok S, Sninsky J, Perkins

Busch MP. HIV and blood transfusions: focus on seroconversion. *Vox Sang.* 1994; 67 Suppl

Campbell ML, Gregory AM & Mauerhan DR (1997). Collection of surgical specimens in total joint arthroplasty. Is routine pathology cost effective? *J Arthroplasty* 1997; 12:60-63. Centers for Disease Control (CDC 1981). Pneumocystis pneumonia--Los Angeles. *MMWR*

Centers for Disease Control (CDC 1982). Update on acquired immune deficiency syndrome

Centers for Disease Control (CDC). Transmission of HIV through bone transplantation: case report and public health recommendations. *MMWR Morb Mortal Wkly Rep.*1988

CDC, no authors listed. Leads from the MMWR. Transmission of HIV through bone

Clark C.R. &Bauer T. Routine pathological examination of operative specimens from

Conrad EU, Gretch DR, Obermeyer KR, Moogk MS, Sayers M, Wilson JJ & Strong DM

Deijkers RL, Bloem RM, Petit PL, Brand R, Vehmeyer SB & Veen MR (1997). Contamination

DiCarlo E.F, Bullough P.G , Steiner G, Bansal M & Kambolis C (1994). Pathological

Eggen BM & Nordbø SA. Transmission of HCV by organ transplantation*. N Engl J Med.* 1992

examination of the femoral head. *Mod Pathol* 1994; 7(1).: 6A.

(AIDS).-United States.*MMWR Morb Mortal Wkly Rep*.1982 Sep 24; 31(37).:507-8,513-4

transplantation: case report and public health recommendations. *JAMA*. 1988 Nov

primary total hip and total knee replacement: another look. *J Bone Joint Surg*

(1995). Transmission of the hepatitis-C virus by tissue transplantation. *J Bone Joint*

of bone allografts: analysis of incidence and predisposing factors. *J Bone Joint Surg*

Aro HT & Aho AJ. Clinical use of bone allografts. *Ann Med*. 1993 Aug;25(4).:403-12.

HG, Sandler SG, Sayers MH, et al. Prevalence of human immunodeficiency virus type 1 p24 antigen in U.S. blood donors--an assessment of the efficacy of testing in donor screening. The HIV-Antigen Study Group. *N Engl J Med.* 1990 Nov

in patients undergoing total hip arthroplasty: A report of two cases. *J Bone Joint*

HA & Vyas GN (1991). Evaluation of screened blood donations for human immunodeficiency virus type 1 infection by culture and DNA amplification of

attention in order to further improve the bone banking process in the near future.

**7. References**

8;323(19).:1312-7.

*Surg* 2000; 82A : 1536-9

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pooled cells. *N Engl J Med.* 1991 Jul 4; 325(1).:1-5.

*Morb Mortal Wkly Rep.* 1981 Jun 5; 30(21).:250-2.

received a femoral-head bone allograft that had been stored by freezing. The graft was procured from a Rhesus positive donor, and the recipient had no other sources of sensitization (Johnson 1985). The immunogenic reaction of allografts is well known and extensively described in literature (Stevenson & Horowitz 1992). This immunogenic response is a reaction on the blood and bone marrow in the allograft. Fresh allografts, which have not been frozen, cause a massive vascular reaction, as has been recently proven by a CAM model (Holzmann et al. 2010). However, freezing of the femoral heads to – 80°C for only three days caused a significant reduction of early vascularisation. Keeping the allografts frozen for longer than one month minimizes the angiogenic potential. Therefore a transfusion reaction after transplantation is unlikely. However, as shown by that case report, sensitization is possible, particularly to the Rhesus (D) (Rh-positive) antigen, which is highly immunogenic. We currently record blood type of all donors, however we only provide Rhesus-compatible grafts to Rhesus-negative women of child-bearing age, in an attempt to prevent problems with future pregnancies or transfusions.

#### **5.4 Audit of a bone bank; further improvements**

As a tool for quality control we performed an intern audit of our hospital bone bank, containing only femoral head allografts from living donors. For this audit we assessed all data from the bone bank registry from November 1994 and March 2010. We also included data from potential allografts which eventually pointed out to be not suitable for transplantation as determined by the aforementioned in-and exclusion bone banking criteria. We retrieved 643 femoral heads as potential allografts from 550 donors. Of 643 harvested femoral heads 242 (38%) were discarded. Based on one or more exclusion criteria 123 grafts were excluded based on the questionnaire or due to incomplete pre-operative donor data or tests. Furthermore, 34 grafts were discarded based on positive microbiological, histopathological or serological examination. In total, 64 grafts were discarded due to missing microbiological, histopathological or serological test after at least 6 months. The rest had to be excluded because of tears in the package, loosening of labels, discovery of malignancies in the donor patient and deceased donors in which serological examination could not take place. We calculated the costs associated with complete testing of one femoral head as potential graft which includes all laboratory, histopathological and bacteriological tests. If all completely tested femoral head allografts would be suitable for donation this bone bank would be financially advantageous, even with routinely performed histopathological assessment. It is never possible that all potential donor allografts are suitable for bone banking. However in our bone bank the major loss of potential allografts is mainly due to managing, administrative and logistic omissions. Therefore currently managing our own hospital bone bank offered no financial benefits. We provide safe and reliable allografts with good accessibility. We calculated that hospital bone banking can be a financially viable strategy, when logistic procedures are more accurate. We made improvements in the logistic procedure of testing and retesting and expect future improvements of our financial bone banking balance.

#### **6. Conclusions**

There are no uniform guidelines for management of a bone bank. The bone bank protocol should meet national law. The described bone bank protocol from our hospital provides for safe and easy accessible allografts. We routinely perform histopathological screening, this practice is extensively discussed on in literature. At this moment we have no financially viable bone bank. This is due to organisational and logistical problems, which have our attention in order to further improve the bone banking process in the near future.

#### **7. References**

262 Wide Spectra of Quality Control

received a femoral-head bone allograft that had been stored by freezing. The graft was procured from a Rhesus positive donor, and the recipient had no other sources of sensitization (Johnson 1985). The immunogenic reaction of allografts is well known and extensively described in literature (Stevenson & Horowitz 1992). This immunogenic response is a reaction on the blood and bone marrow in the allograft. Fresh allografts, which have not been frozen, cause a massive vascular reaction, as has been recently proven by a CAM model (Holzmann et al. 2010). However, freezing of the femoral heads to – 80°C for only three days caused a significant reduction of early vascularisation. Keeping the allografts frozen for longer than one month minimizes the angiogenic potential. Therefore a transfusion reaction after transplantation is unlikely. However, as shown by that case report, sensitization is possible, particularly to the Rhesus (D) (Rh-positive) antigen, which is highly immunogenic. We currently record blood type of all donors, however we only provide Rhesus-compatible grafts to Rhesus-negative women of child-bearing age, in an attempt to

As a tool for quality control we performed an intern audit of our hospital bone bank, containing only femoral head allografts from living donors. For this audit we assessed all data from the bone bank registry from November 1994 and March 2010. We also included data from potential allografts which eventually pointed out to be not suitable for transplantation as determined by the aforementioned in-and exclusion bone banking criteria. We retrieved 643 femoral heads as potential allografts from 550 donors. Of 643 harvested femoral heads 242 (38%) were discarded. Based on one or more exclusion criteria 123 grafts were excluded based on the questionnaire or due to incomplete pre-operative donor data or tests. Furthermore, 34 grafts were discarded based on positive microbiological, histopathological or serological examination. In total, 64 grafts were discarded due to missing microbiological, histopathological or serological test after at least 6 months. The rest had to be excluded because of tears in the package, loosening of labels, discovery of malignancies in the donor patient and deceased donors in which serological examination could not take place. We calculated the costs associated with complete testing of one femoral head as potential graft which includes all laboratory, histopathological and bacteriological tests. If all completely tested femoral head allografts would be suitable for donation this bone bank would be financially advantageous, even with routinely performed histopathological assessment. It is never possible that all potential donor allografts are suitable for bone banking. However in our bone bank the major loss of potential allografts is mainly due to managing, administrative and logistic omissions. Therefore currently managing our own hospital bone bank offered no financial benefits. We provide safe and reliable allografts with good accessibility. We calculated that hospital bone banking can be a financially viable strategy, when logistic procedures are more accurate. We made improvements in the logistic procedure of testing and retesting and expect future

There are no uniform guidelines for management of a bone bank. The bone bank protocol should meet national law. The described bone bank protocol from our hospital provides for safe and easy accessible allografts. We routinely perform histopathological screening, this

prevent problems with future pregnancies or transfusions.

**5.4 Audit of a bone bank; further improvements**

improvements of our financial bone banking balance.

**6. Conclusions**

Quality Control in Hospital Bone Banking 265

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**15**

Alphus Dan Wilson

*United States of America*

**Future Applications of Electronic-Nose**

*USDA Forest Service, Southern Hardwoods Laboratory*

**Technologies in Healthcare and Biomedicine**

The development and utilization of many new electronic-nose (e-nose) applications in the healthcare and biomedical fields have continued to rapidly accelerate over the past 20 years. Innovative e-nose technologies are providing unique solutions to a diversity of complex problems in biomedicine that are now coming to fruition. A wide range of electronic-nose instrument types, based on different operating principles and mechanisms, has facilitated the creation of different types and categories of medical applications that take advantage of the unique strengths and advantages of specific sensor types and sensor arrays of different individual instruments. Electronic-nose applications have been developed for a wide range of healthcare sectors including diagnostics, immunology, pathology, patient recovery, pharmacology, physical therapy, physiology, preventative medicine, remote healthcare, and wound and graft healing. E-nose biomedical applications range from biochemical testing, blood compatibility, disease diagnoses, drug purity, monitoring metabolic levels, organ dysfunction, and telemedicine. This review summarizes some of the key technological developments of electronic-nose technologies, arising from past and recent biomedical research, and identifies a variety of future e-nose applications currently under development which have great potential to advance the effectiveness and efficiency of biomedical treatments and healthcare services for many years. A concise synthesis of the major electronic-nose technologies developed for healthcare and medical applications since the 1980s is provided along with a detailed assessment and analysis of future potential advances in electronic aroma detection (EAD) technologies that will provide effective solutions to newly-emerging problems in the healthcare industry. These new e-nose solutions will provide greatly improved quality controls for healthcare decisions and diagnoses as well as badly needed final confirmations of appropriate patient treatments. The purpose of this chapter is to provide some detailed insights into current and future e-nose applications that will yield a variety of new solutions to detection-related tasks and difficult problems in the fields of healthcare and biomedicine. The uses of electronic-noses for quality control (QC) and quality assurance (QA) issues, associated with numerous diagnostic-

testing activities conducted within the medical field, also are discussed.

**2. History of Electronic-nose developments for biomedical applications**

Use of the olfactory sense (of smell) as an indicator of disease probably originated with Hippocrates around 400 BC. Observations that unusual human odors or aromas provided

**1. Introduction**

### **Future Applications of Electronic-Nose Technologies in Healthcare and Biomedicine**

Alphus Dan Wilson

*USDA Forest Service, Southern Hardwoods Laboratory United States of America*

#### **1. Introduction**

266 Wide Spectra of Quality Control

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Veen MR, Bloem RM & Petit PL.(1994).Sensitivity and negative predictive value of swab

Vehmeyer SB, Slooff AR, Bloem RM & Petit PL (2002). Bacterial contamination of femoral head allografts from living donors. *Acta Orthop Scand*. 2002 Apr;73(2).:165-9. Zwitser EW, de Gast A, Basie MJ, van Kemenade FJ & van Royen BJ (2009). B-cell lymphoma

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bank bone*. J Bone Joint Surg Am* 1954, 36-A: 160-162.

lumbar spine fusion*. J Bone Joint Surg Br* 71:677–680

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(2009). Acetabular revision with impacted morselized cancellous bone graft and a cemented cup in patients with rheumatoid arthritis. A concise follow-up, at eight to nineteen years, of a previous report. *J Bone Joint Surg Am.* 2009 Mar 1; 91(3).:646-51. Schreurs BW, Keurentjes JC, Gardeniers JW, Verdonschot N, Slooff TJ & Veth

RP(2009).Acetabular revision with impacted morselized cancellous bone grafting and a cemented acetabular component: a 20- to 25-year follow-up. *J Bone Joint Surg*

femoral impaction grafting in revision total hip arthroplasty. *J Bone Joint Surg Am*.

and femoral reconstruction with impacted graft and cement*. Clin Orthop Relat Res*

of acetabular reconstruction with bone impaction grafting in dysplastic hips. *Clin*

Histopathology of retrieved allografts of the femoral head. *J Bone Joint Surg Br*

Sukegawa K, Okada T & Fukuda M (2011). Surgical treatment of scoliosis using allograft bone from a regional bone bank. *Arch Orthop Trauma Surg*. 2011 Feb;

to the process of new bone formation by induction. *Am J Surg*. 1953 Mar;85(3).:444-9

cultures in musculoskeletal allograft procurement. *Clin Orthop Relat Res.* 1994

in retrieved femoral heads: a long term follow up. *BMC Musculoskelet Disord* 2009;

orthopaedic bone bank in the Netherlands*. Cell Tissue Bank*. 2010 Nov 13. [Epub

The development and utilization of many new electronic-nose (e-nose) applications in the healthcare and biomedical fields have continued to rapidly accelerate over the past 20 years. Innovative e-nose technologies are providing unique solutions to a diversity of complex problems in biomedicine that are now coming to fruition. A wide range of electronic-nose instrument types, based on different operating principles and mechanisms, has facilitated the creation of different types and categories of medical applications that take advantage of the unique strengths and advantages of specific sensor types and sensor arrays of different individual instruments. Electronic-nose applications have been developed for a wide range of healthcare sectors including diagnostics, immunology, pathology, patient recovery, pharmacology, physical therapy, physiology, preventative medicine, remote healthcare, and wound and graft healing. E-nose biomedical applications range from biochemical testing, blood compatibility, disease diagnoses, drug purity, monitoring metabolic levels, organ dysfunction, and telemedicine. This review summarizes some of the key technological developments of electronic-nose technologies, arising from past and recent biomedical research, and identifies a variety of future e-nose applications currently under development which have great potential to advance the effectiveness and efficiency of biomedical treatments and healthcare services for many years. A concise synthesis of the major electronic-nose technologies developed for healthcare and medical applications since the 1980s is provided along with a detailed assessment and analysis of future potential advances in electronic aroma detection (EAD) technologies that will provide effective solutions to newly-emerging problems in the healthcare industry. These new e-nose solutions will provide greatly improved quality controls for healthcare decisions and diagnoses as well as badly needed final confirmations of appropriate patient treatments. The purpose of this chapter is to provide some detailed insights into current and future e-nose applications that will yield a variety of new solutions to detection-related tasks and difficult problems in the fields of healthcare and biomedicine. The uses of electronic-noses for quality control (QC) and quality assurance (QA) issues, associated with numerous diagnostictesting activities conducted within the medical field, also are discussed.

#### **2. History of Electronic-nose developments for biomedical applications**

Use of the olfactory sense (of smell) as an indicator of disease probably originated with Hippocrates around 400 BC. Observations that unusual human odors or aromas provided

Future Applications of Electronic-Nose Technologies in Healthcare and Biomedicine 269

odors associated with completely different diseases. This occurred because different diseases can result in the production of very similar compounds even though the mechanism of disease is quite different. In other cases such as for use of the term "fishy" for hypermethioninemia and uremia, both of these diseases cause the buildup of similar or identical compounds in the blood due to similar physiological processes that are often referred to as in-born genetic or metabolic diseases resulting from the absence of certain enzymes or the failure of certain organs. Many other metabolic diseases caused by genetic enzyme deficiencies are associated with various distinctive odors due to the accumulation of

Some descriptive aromas, such as maple syrup and pox stench, are so diagnostic that the aroma was named after the specific disease referred to by name. Other diagnostic terms for descriptive aromas include fetor hepaticus, diabetic breath, and uremic breath which have been included in common medical vocabulary and continue to be used to some extent even in contemporary vernacular. Once modern analytical instrumentation became available in the twentieth century, the actual volatile compounds responsible for these characteristic smells began to be identified. Probably the first such identification was done by Linus Pauling, the noted chemist who was able to freeze out and identify some of the volatiles in urine using cold traps, followed by gas chromatography (Pauling et al., 1971). Many other discoveries of VOCs associated with specific human smells related to particular diseases followed in subsequent years leading up to the identification of diagnostic bioindicators of disease. These compounds are highly correlated with the presence of specific diseases in the

The discovery and recognition of particular volatile organic compounds (VOCs), released from various diseased human body parts or fluids derived from these tissues, have been found to be associated with specific human diseases through the use of specialized modern analytical instruments. These instruments have included such analytical machines as gas chromatographs working in tandem with mass spectrometers (GC-MS) and other such technical instruments used in analytical chemistry. The results of intense chemical analyses from numerous research studies have been the identification of many volatile biomarkers of disease and their associated chemical structures. The identification of unique molecular markers (volatile metabolites) associated with particular diseases has become an extremely effective and powerful tool for the early detection of diseased tissues and infectious agents in the human body. For example, the analysis of patients' breath odors has had a long history of application for the detection of various human diseases, not only respiratory diseases. Even though the human breath contains hundreds of volatile organic compounds at low concentrations, relatively few (less than fifty) of these are detected in the majority of healthy humans under normal physiological conditions (Phillips et al., 1999a). However, a much smaller number of aberrant VOCs are often found only in patients when disease is present somewhere in their bodies. Thus, the association of specific volatile metabolites, released within the expired human breath of patients, not only provides indicators of particular diseases, but also reflect the overall physiological state as an indication of general health and a useful index of disease (Jacoby, 2004). These volatile markers of disease often are released several hours to several days before outwardly-noticeable physical symptoms of illness appear and thus provide early indicators of disease or physiological disorders. New molecular markers that are indicators of specific diseases, both infectious and

undecomposed metabolites in the body.

body as discussed in the following section.

**2.2 Discovery of bioindicators of disease**

some indication of human ailments caused early medical practitioners to recognize that the presence of human diseases changed the odor of bodily excretions that could be used to diagnose certain common diseases.

#### **2.1 Early use of aroma-detection in evaluating health conditions**

Medical doctors have utilized the sense of smell to facilitate determinations of the physical state and general health of their patients for centuries. The application of smell as useful sensory clues used by physicians to identify the causes of human ailments resulted in the development of qualitatively descriptive odors (or aromas) and specialized terms used to describe and identify odors associated with specific human diseases and physiological disorders. Some descriptive aromas found to be associated with some common human diseases are presented in Table 1. The use of olfactory information provided valuable additional information for physicians in assessing patient conditions and formulating accurate diagnoses before modern analytical equipment and chemical-detection devices became available for this purpose. Notice that in some cases the same term, such as "aminelike" for bacterial vaginosis and bladder infections, occasionally was used to describe common

Table 1. Descriptive aromas previously used for diagnosing human diseases

odors associated with completely different diseases. This occurred because different diseases can result in the production of very similar compounds even though the mechanism of disease is quite different. In other cases such as for use of the term "fishy" for hypermethioninemia and uremia, both of these diseases cause the buildup of similar or identical compounds in the blood due to similar physiological processes that are often referred to as in-born genetic or metabolic diseases resulting from the absence of certain enzymes or the failure of certain organs. Many other metabolic diseases caused by genetic enzyme deficiencies are associated with various distinctive odors due to the accumulation of undecomposed metabolites in the body.

Some descriptive aromas, such as maple syrup and pox stench, are so diagnostic that the aroma was named after the specific disease referred to by name. Other diagnostic terms for descriptive aromas include fetor hepaticus, diabetic breath, and uremic breath which have been included in common medical vocabulary and continue to be used to some extent even in contemporary vernacular. Once modern analytical instrumentation became available in the twentieth century, the actual volatile compounds responsible for these characteristic smells began to be identified. Probably the first such identification was done by Linus Pauling, the noted chemist who was able to freeze out and identify some of the volatiles in urine using cold traps, followed by gas chromatography (Pauling et al., 1971). Many other discoveries of VOCs associated with specific human smells related to particular diseases followed in subsequent years leading up to the identification of diagnostic bioindicators of disease. These compounds are highly correlated with the presence of specific diseases in the body as discussed in the following section.

#### **2.2 Discovery of bioindicators of disease**

268 Wide Spectra of Quality Control

some indication of human ailments caused early medical practitioners to recognize that the presence of human diseases changed the odor of bodily excretions that could be used to

Medical doctors have utilized the sense of smell to facilitate determinations of the physical state and general health of their patients for centuries. The application of smell as useful sensory clues used by physicians to identify the causes of human ailments resulted in the development of qualitatively descriptive odors (or aromas) and specialized terms used to describe and identify odors associated with specific human diseases and physiological disorders. Some descriptive aromas found to be associated with some common human diseases are presented in Table 1. The use of olfactory information provided valuable additional information for physicians in assessing patient conditions and formulating accurate diagnoses before modern analytical equipment and chemical-detection devices became available for this purpose. Notice that in some cases the same term, such as "aminelike" for bacterial vaginosis and bladder infections, occasionally was used to describe common

**Disease / Disorder Body source Descriptive aroma References**

Fetor hepaticus Breath Newly-mown clover Hayden, 1980 Gout Skin Gouty odor Liddell, 1976 Hyperaminoaciduria Infant skin Dried malt or hops Liddell, 1976

Ketoacidosis Breath Acetone-like Hayden, 1980

Scrofula Body Stale beer Liddell, 1976 Smallpox Skin Pox stench Liddell, 1976

Uremia Breath Fishy, ammonia Hayden, 1980

Table 1. Descriptive aromas previously used for diagnosing human diseases

Anaerobic infection Skin, sweat Rotten apples Pavlou & Turner, 2000 Bacterial vaginosis Vaginal fluid Amine-like Pavlou & Turner, 2000 Bladder infection Urine Amine-like Pavlou & Turner, 2000

Hypermethioninemia Infant breath Sweet, fruity, fishy Liddell, 1976; Hayden,

Isovaleric acidemia Skin, breath Sweaty, cheesy Hayden, 1980; Pavlou

Liver failure Breath Musty fish, feculent Hayden, 1980; Smith,

Maple syrup disease Sweat, urine Maple syrup Liddell, 1976; Pavlou

*Pseudomonas* infection Skin, sweat Grape Pavlou & Turner, 2000

Trimethylaminuria Skin, urine Fishy Pavlou & Turner, 2000 Typhoid Skin Freshly-baked bread Liddell, 1976; Hayden,

Yellow fever Skin Butcher's shop Liddell, 1976; Hayden,

Heart Dimethyl sulfide Smith, 1982

1980

1982

1980

1980

& Turner, 2000

& Turner, 2000

**2.1 Early use of aroma-detection in evaluating health conditions**

diagnose certain common diseases.

Congestive heart

failure

The discovery and recognition of particular volatile organic compounds (VOCs), released from various diseased human body parts or fluids derived from these tissues, have been found to be associated with specific human diseases through the use of specialized modern analytical instruments. These instruments have included such analytical machines as gas chromatographs working in tandem with mass spectrometers (GC-MS) and other such technical instruments used in analytical chemistry. The results of intense chemical analyses from numerous research studies have been the identification of many volatile biomarkers of disease and their associated chemical structures. The identification of unique molecular markers (volatile metabolites) associated with particular diseases has become an extremely effective and powerful tool for the early detection of diseased tissues and infectious agents in the human body. For example, the analysis of patients' breath odors has had a long history of application for the detection of various human diseases, not only respiratory diseases. Even though the human breath contains hundreds of volatile organic compounds at low concentrations, relatively few (less than fifty) of these are detected in the majority of healthy humans under normal physiological conditions (Phillips et al., 1999a). However, a much smaller number of aberrant VOCs are often found only in patients when disease is present somewhere in their bodies. Thus, the association of specific volatile metabolites, released within the expired human breath of patients, not only provides indicators of particular diseases, but also reflect the overall physiological state as an indication of general health and a useful index of disease (Jacoby, 2004). These volatile markers of disease often are released several hours to several days before outwardly-noticeable physical symptoms of illness appear and thus provide early indicators of disease or physiological disorders. New molecular markers that are indicators of specific diseases, both infectious and

Future Applications of Electronic-Nose Technologies in Healthcare and Biomedicine 271

Analysis of expired human breath is considered particularly valuable because it can be monitored noninvasively (without causing physical damage to patients), yet provide information about the chemical and physiological state of the entire body. The reason that information about the physical health of the entire body is possible by the analysis of expired breath is because most volatile metabolites of infectious agents of disease, or those produced from abnormal tissues, are eventually eliminated from the body through the lungs, often soon after being formed within diseased tissues. Alternatively, other less volatile abnormal metabolites are eliminated through the urine which may be similarly

Cao and Duan (2006) summarized some of the advantages and disadvantages of breath analysis for clinical practice and diagnosis. They found breath tests were noninvasive, easily repeated, and caused less discomfort and embarrassment to patients than blood and urine tests. Breath samples closely reflected arterial concentrations and provided much less complicated mixtures than serum or urine analyses and more direct information on respiratory function than by other means. They listed limitations of breath testing for clinical practice to include the lack of standardization of analytical methods, the high water content of breath samples affecting detection, relatively expensive costs compared to simple chemical tests (but much less time-consuming for results), and the lack of well-established links between breath VOCs and certain kinds of diseases. Biomarkers in chronic obstructive pulmonary disease (COPD) also may be useful in aiding diagnosis, monitoring exacerbations, evaluating effects of drugs, and defining specific phenotypes of disease (Borrill et al., 2008). Frey & Suki (2008) found risk assessments, disease progression, and control of asthma and COPD required multidimensional fluctuation analysis of the dynamics of lung-function parameters that needed to be quantified and monitoring via precise biomarkers of these diseases using instruments capable of direct, electronic monitoring of these biomarkers. The importance of the use of biomarkers for the detection of disease has become so prominent that Bentham Science, a leading international publisher of high quality scientific journals, decided to launch a new journal called *Recent Patents on Biomarkers* in January 2011 to publish reviews and research articles written by experts on recent patents and research relating to biomarkers in basic and applied, medical, environmental, and pharmaceutical research, and including patent biomarker applications, clinical development, and molecular

**3. Current e-nose technologies utilized in healthcare and biomedicine**

Electronic-noses are ideal instruments for biomedical uses because of their versatility, lowcost, rapid output of results, capabilities of continuous operation (for physiologicalmonitoring purposes), and the wide range of VOCs and other cellular chemical constituents that may be analyzed. The potential for miniaturization of electronic-nose devices also is great due to their microcircuitry and microsensor components. Some key ways in which enoses have been particularly useful in various sectors of the healthcare industry are

A variety of different types of e-noses, based on different working principles, have been used for biomedical tasks including conductive polymers (CP), metal-oxide semiconductor (MOS), quartz crystal microbalance (QCM), and surface acoustic waves (SAW) among

analyzed using aroma-sensing instruments such as electronic noses.

diagnostics.

discussed in the following sections.

**3.1 Electronic-nose technology types and applications**

noninfectious, are being increasingly revealed by new scientific research. Some examples of these volatile molecular biomarkers (or bioindicators) of disease and physiological disorders, reported hitherto by various researchers, are summarized in Table 2.

Table 2. Molecular biomarker VOCs of specific human diseases and disorders

noninfectious, are being increasingly revealed by new scientific research. Some examples of these volatile molecular biomarkers (or bioindicators) of disease and physiological

Garner et al., 2009

Carpagnano et al., 2003; Phillips et al., 2004

Crofford et al., 1997; Ping et al. 1997; Novak

Kaizu, 1976; Van den Velde et al., 2009

Bondy & Rosenberg,

Manolis, 1983; Gordon et al., 1985; Preti et al., 1988; Phillips et al., 1999b, 2003a

Bondy & Rosenberg,

Smith et al., 1969; Phillips et al., 1993

et al., 2007

1980

1980

2008

1980

1980

disorders, reported hitherto by various researchers, are summarized in Table 2.

 Molecular biomarker VOCs of specific human diseases and disorders

Hepatic encephalopathy 3-methylbutanal Goldberg, 1981

Liver cancer Hexanal, 1-octen-3-ol, octane Xue et al., 2008

2-imidazolelactic acid, 2-imidazoleacetic acid Analysis of expired human breath is considered particularly valuable because it can be monitored noninvasively (without causing physical damage to patients), yet provide information about the chemical and physiological state of the entire body. The reason that information about the physical health of the entire body is possible by the analysis of expired breath is because most volatile metabolites of infectious agents of disease, or those produced from abnormal tissues, are eventually eliminated from the body through the lungs, often soon after being formed within diseased tissues. Alternatively, other less volatile abnormal metabolites are eliminated through the urine which may be similarly analyzed using aroma-sensing instruments such as electronic noses.

Cao and Duan (2006) summarized some of the advantages and disadvantages of breath analysis for clinical practice and diagnosis. They found breath tests were noninvasive, easily repeated, and caused less discomfort and embarrassment to patients than blood and urine tests. Breath samples closely reflected arterial concentrations and provided much less complicated mixtures than serum or urine analyses and more direct information on respiratory function than by other means. They listed limitations of breath testing for clinical practice to include the lack of standardization of analytical methods, the high water content of breath samples affecting detection, relatively expensive costs compared to simple chemical tests (but much less time-consuming for results), and the lack of well-established links between breath VOCs and certain kinds of diseases. Biomarkers in chronic obstructive pulmonary disease (COPD) also may be useful in aiding diagnosis, monitoring exacerbations, evaluating effects of drugs, and defining specific phenotypes of disease (Borrill et al., 2008).

Frey & Suki (2008) found risk assessments, disease progression, and control of asthma and COPD required multidimensional fluctuation analysis of the dynamics of lung-function parameters that needed to be quantified and monitoring via precise biomarkers of these diseases using instruments capable of direct, electronic monitoring of these biomarkers.

The importance of the use of biomarkers for the detection of disease has become so prominent that Bentham Science, a leading international publisher of high quality scientific journals, decided to launch a new journal called *Recent Patents on Biomarkers* in January 2011 to publish reviews and research articles written by experts on recent patents and research relating to biomarkers in basic and applied, medical, environmental, and pharmaceutical research, and including patent biomarker applications, clinical development, and molecular diagnostics.

#### **3. Current e-nose technologies utilized in healthcare and biomedicine**

Electronic-noses are ideal instruments for biomedical uses because of their versatility, lowcost, rapid output of results, capabilities of continuous operation (for physiologicalmonitoring purposes), and the wide range of VOCs and other cellular chemical constituents that may be analyzed. The potential for miniaturization of electronic-nose devices also is great due to their microcircuitry and microsensor components. Some key ways in which enoses have been particularly useful in various sectors of the healthcare industry are discussed in the following sections.

#### **3.1 Electronic-nose technology types and applications**

A variety of different types of e-noses, based on different working principles, have been used for biomedical tasks including conductive polymers (CP), metal-oxide semiconductor (MOS), quartz crystal microbalance (QCM), and surface acoustic waves (SAW) among

Future Applications of Electronic-Nose Technologies in Healthcare and Biomedicine 273

**E-nose**

Experimental model

Fox 4000 Flavor

Cyranose 320 Identify

PEN 2 Clinical

unspecified Diagnosis of

unspecified Detect chronic

Cyranose 320 Discriminate

NST 3320 Diagnosis of

JPL ENose Detect &

experimental model

model

EN

Table 3. Electronic-nose uses in hospitals and universities around the world

Bloodhound

DE 101 Detect renal

**utilized Application References**

Thaler et al.,

Zhu et al., 2004

Dutta et al., 2005

Voss et al., 2005

2005

2006

Persaud, 2006

Anthes, 2008

Dent, 2010

Kateb et al., 2009

Lazar et al., 2010

Montuschi, 2010

Kolk et al., 2010

Humphreys et al., 2011

Erzurum et al.,

Moens et al.,

2000

Distinguish cerebrospinal

analysis for drug formulation

*Staphalococcus*

dysfunction

diagnoses of bacteria

Burn and wound infection types

diseases via breath

differentiate brain cancers

lung disease

inflammation airway diseases

Asthma detection

Diagnosis of Tuberculosis

ventilatorassociated pneumonia

cancer

fluid

**Country Hospital, University**

USA University of

USA Merck Research

Germany University of

Belgium University of

USA University of

USA California Institute

Australia Prince Charles

Netherlands Amsterdam

United Kingdom

United Kingdom

United Kingdom **or Research Facility**

Pennsylvania

Laboratories

Birmingham

Antwerp

Heartlands Hospital

USA Cleveland Clinic unspecified Diagnose lung

Applied Sciences

South Manchester University Hospital

Pennsylvania

of Technology

Academic Medical

Italy Catholic University experimental

Medical Research

Gloucestershire Royal Hospital

Hospital

Center

Tanzania National Institute of

others. Each e-nose technology has different advantages, disadvantages, and limitations that largely determine what types of medical applications that individual e-nose sensor types are best suited for in practical clinical settings.

#### **3.2 Point-of-care medicine**