dental trauma is one of the most painful and stressful injuries in young patients and requires both expedient and expert management by a dental practitioner .
children are always keen to explore their surroundings without realizing the risks they are exposed to .
the world health organization has defined exarticulation as the complete displacement of a tooth from its alveolar socket due to traumatic injury . in permanent dentition
, it may occur as the result of fights or sport injuries , while falls against hard objects are among the frequent causes in the primary dentition .
research has shown that an exarticulated tooth can be re - planted without complications if it is reinserted into the socket within 20 minutes when stored dry , and within one to three hours if placed in a suitable storage medium . when the tooth is kept dry for more than 20 minutes ,
its pdl cells begin to necrotize , and upon replantation , inflammation and resorption develop in proportion to the extra - oral dry time . many storage media such as milk , saliva , hank s balanced salt solution ( hbss ) ,
, hbss has been proposed as the storage medium of choice for treatment of avulsed teeth by the american associations of endodontists .
however , the major disadvantage of hbss and many of the other aforementioned media is that they are not easily available at places where these traumatic injuries occur .
hence , there is a need to identify a storage medium that will be readily available , yet effective .
aloe vera has recently gained popularity in the medicinal field because of its antidiabetic , anticancer , and antibiotic properties .
its anti - inflammatory properties enhance wound healing whether it is used topically or orally .
because of its increased use , the plant can be found commonly in most households , nurseries , parks and recreation areas .
also , several reputable suppliers have produced a stabilized aloe vera gel for use alone or in combined formulations [ 68 ] .
the purpose of this study was to assess the efficacy of three easily available storage media , milk , aloe vera , and egg white , in preserving the viability of pdl cells of avulsed teeth in vitro .
forty - five non - carious human premolar teeth with normal periodontium and closed apices , undergoing extraction for orthodontic therapeutic purposes , were included in this study ( fig .
they were divided into three equal groups using simple random allocation ( using a chit  pull system ) . 

group i : stored in milk ( control group)group ii : stored in aloe vera ( experimental group)group iii : stored in egg white ( experimental group ) .
group i : stored in milk ( control group ) group ii : stored in aloe vera ( experimental group ) group iii : stored in egg white ( experimental group ) .
non - carious mature premolar teeth were extracted pasteurized , homogenized milk with fat content of 3gm/100ml ( saras dairy ) at room temperature was used as the control group . to prepare aloe vera gel ,
a kitchen knife was used to cut off the outer spiked edges of the leaves . after removing the outer skin of the leaves ,
the contents were blended thoroughly and filtered through a piece of muslin cloth and were placed into a glass jar with a tight fitting lid   ( fig .
egg white was separated from the yolk and collected in a bowl using the half shelves .
following extractions , the coronal third of pdl was scraped with a curette to remove cells likely to be damaged due to instrumentation . to simulate avulsion injury ,
teeth were kept in mud for 15 minutes immediately after removal of the damaged cells from the coronal one - third   ( fig .
after 30 minutes , the root surface was irrigated twice with sterile isotonic saline to remove the residual storage media .
the apical two - third of the root surfaces ( as measured from epithelial attachment ) were scraped in a petri dish to obtain the pdl cells .
preparation of aloe vera gel mud for 15 minutes to simulate avulsion injury the obtained scrapings were subsequently added to a falcon tube containing 2.5 ml of phosphate buffer   ( fig .
4 ) . to the above mixture , 0.5 mg of type i collagenase was added , and this mixture was incubated for 30 minutes . following incubation ,
pdl cells ( from apical two - third of the root surface ) were scraped in a petri dish the supernatant was discarded and the centrifuged residue was collected . to this , an equal volume of 0.4% trypan blue stain was added , and it was mixed well .
trypan blue stains non - viable cells blue and viable cells appear colorless or pink . following staining ,
the number of cells ( total and non - viable ) was determined by counting the cells overlying a 41 mm area of the counting chamber ( figs . 5 and 6 ) .
microscopic view of viable and non - viable cells in milk microscopic view of viable and non - viable cells in aloe vera the viable cell percentage was calculated as : [ ( total cells - stained cells)/total cells ] 100 the mean and standard deviation of the number of viable cells from different storage media were calculated and statistical analysis was performed .
inter - group comparison was done using one - way analysis of variance ( anova ) complemented by the tukey s test .
the number of cells ( total and non - viable ) was determined by counting the cells overlying a 41 mm area of the counting chamber ( figs . 5 and 6 ) .
microscopic view of viable and non - viable cells in milk microscopic view of viable and non - viable cells in aloe vera the viable cell percentage was calculated as : [ ( total cells - stained cells)/total cells ] 100 the mean and standard deviation of the number of viable cells from different storage media were calculated and statistical analysis was performed .
inter - group comparison was done using one - way analysis of variance ( anova ) complemented by the tukey s test .
the teeth stored in aloe vera demonstrated significantly higher percentage of viable pdl cells ( 87.3% ) compared to those stored in egg white ( 75.7% ) or milk ( 74.0% ) ( table 1 , graph 1 ) .
aloe vera showed higher number of viable cells when compared to milk and egg white ( p=0.0001 between groups i  ii and ii  iii , p=0.4757 between groups i  iii , table 2 ) .
the mean values of viable and nonviable and percentage of viable pdl cell count in the study groups and control group percentage of viable cells inter - group comparison using one - way anova complemented by the tukey s test .
post hoc power analysis was done using g power software 3.0.10 . by fixing the alpha at 0.05 ,
the calculated effect size from the given values was 0.55 and the power of the study was 0.9 or 90% .
recent clinical studies have shown that avulsed teeth replanted within five minutes have the best prognosis , resulting in much higher reattachment success . to avoid root resorption and maintain pdl cell viability ,
various transport media have been proposed to store the tooth in , if immediate re - plantation is not possible .
the basic principle behind the use of these media is to transport the tooth to an environment similar to that of the oral cavity .
various studies have found hbss , viaspan , and eagle s medium to be suitable for transportation of avulsed teeth as they preserved pdl cell viability . however , the main disadvantages of these media are high cost and lack of availability
. therefore , there should be an effort to find a storage medium that is easily available and cheap .
hence , the present study was carried out to compare the effectiveness of two of the most commonly available and cost effective storage media , aloe vera and egg white .
therefore , noncarious mature human premolar teeth undergoing extraction for orthodontic therapeutic purposes were selected .
for the same reason , teeth from young healthy individuals without periodontal disease were included in the study .
on average it takes about 10 to 15 minutes for the victim and the person caring for him / her to recover from the traumatic event and act appropriately .
according to pohl et al , pdl cells remain in a non - compromised state with up to 15 minutes of dry time .
other investigators have shown that at two hours of dry time , no vital pdl cells remain . in the present study , it was decided to keep the extracted tooth in mud for at least 15 minutes to simulate avulsion injury , and to ensure that sufficient viable pdl cells are available for assessment . following a 15-minute dry time , the teeth were placed in different storage media for 30 minutes .
this period was very important because the pdl cells were most susceptible to damage during this time .
it is a physiological medium , which contains fewer bacteria compared to saliva ; therefore it is one of the most frequently used transport media . due to these advantages , in the present study , milk was chosen as the positive control . in the present study ,
aloe vera gel was chosen as a transport medium because it contains 99% water and over 75 nutrients , which include 20 minerals , 19 amino acids and 12 vitamins .
the human body requires 22 amino acids to maintain good health , eight of which are essential , as the body can not synthesize them .
all of these eight essential amino acids and 11 of 14 secondary amino acids are found in aloe vera .
it is thought to promote healing ; therefore it can be used in surgical wounds , in root canal treatment as an analgesic dressing , and around dental implants to control inflammation . in a recent study , human kidney cell death rate
egg white from a single egg contains 4.7 grams of 40 different proteins , 0.3 grams of carbohydrate , 62 milligrams of sodium , and the remainder being water . a method suggested by ragnarsson et al .
has been quoted in the literature to evaluate the efficacy of different storage media in preserving the viability of dental fibroblasts .
ragnarsson et al . removed fibroblasts from the root surfaces and cultured them after adding them to a storage medium .
however , doyle et al . placed the extracted teeth directly in a storage medium . at a pre - determined time , the teeth were taken out of the medium and pdl cells were isolated to evaluate cell viability . since the method by doyle et al .
was found to closely replicate the clinical scenario , it was followed in this study . to quantitate the number of viable pdl cells in the current study and to preserve maximum cell viability ,
the root surfaces were treated with type i collagenase for rapid cell retrieval and cellular integrity .
trypan blue exclusion assay quickly and easily differentiates non - viable cells from viable ones . as chromophore present on cell membranes
is negatively charged , it does not take up trypan blue stain unless the membrane is damaged ; consequently all the viable cells are excluded as they do not pick up the dye .
trypan blue stain used in this study assessed only viability of the cells and not the actual physiologic health and metabolic capabilities of them . according to our results , maximum percentage of viable cells
was found in aloe vera ( 87.3% ) , followed by egg white ( 75.7% ) and milk ( 74.07% ) .
aloe vera maintained the highest number of viable cells , which could be attributed to its composition .
aloe vera also contains allantoin , which has been found to stimulate fibroblast activity and collagen proliferation . in a recent study ,
green tea extract also showed higher number of viable cells compared to milk , which was used as a positive control group .
egg white maintained less viable cells than aloe vera , which may be attributed to its high ph ( 9.38 ) , and also to the large amount of proteins in egg white , which might act as foreign bodies . in our study , milk showed the least number of viable cells ( 74.0% ) , which could be attributed to the presence of various enzymes in the milk , which could be harmful to the fibroblasts of the pdl .
found that milk was unable to regenerate depleted cell metabolites and restore the viability of pdl cells .
also , because of the lack of a cell energy source and ions in milk , repopulation of pdl was not permitted .
milk was found to be a compatible storage medium only when it was cold and fresh .
immediate re - implantation is the best treatment for exarticulated teeth , provided that the teeth have viable pdl cells at the time of re - implantation .
if that is not possible , storing the tooth in a storage medium would be the next best option to preserve its viability until re - implantation is possible . within the limitations of this study
, it appears that aloe vera maintains pdl cell viability better than egg white or milk . because of its superiority as a storage medium , and other medicinal properties ,
it is recommended that it should be made easily available in the form of gel or other formulations over the counter , and be added to first aid kits at schools and sport clubs , so that it can be easily accessed at the time of trauma / injury to teeth .
further studies , including in vivo studies , are needed to investigate whether aloe vera can maintain pdl cell viability if maintained at lower temperatures or at extended extra oral dry time .
 objectives : this study was undertaken to evaluate the viability of periodontal ligament ( pdl ) cells of avulsed teeth in three different storage media.materials and methods : forty - five premolars extracted for orthodontic therapeutic purposes were randomly and equally divided into three groups based on storage media used [ group i : milk ( control ) ; group ii : aloe vera ( experimental ) ; group iii : egg white ( experimental ) ] . following extractions , the teeth were placed in one of the three different storage media for 30 minutes , following which the scrapings of the pdl from these teeth were collected in falcon tubes containing collagenase enzyme in 2.5 ml of phosphate buffered saline . 
 the tubes were subsequently incubated for 30 minutes and centrifuged for five minutes at 800 rpm . 
 the obtained pdl cells were stained with trypan blue and were observed under optical microscope . 
 the percentage of viable cells was calculated.results:aloe vera showed the highest percentage of viable cells ( 114.38.0 ) , followed by egg white ( 100.96.3 ) and milk ( 101.17.3).conclusion : within the limitations of this study , it appears that aloe vera maintains pdl cell viability better than egg white or milk . 
INTRODUCTION
MATERIALS AND METHODS
Determination of the number of cells (total and viable):
RESULTS
DISCUSSION
CONCLUSION