some penicillins have been reported to cause occupational diseases , including bronchial asthma ( 1 , 2 ) , anaphylaxis ( 3 ) , contact urticaria ( 4 ) , contact dermatitis ( 5 ) , and hypersensitivity pneumonitis ( 6 ) , in pharmaceutical workers and health care professionals .
occupational contact urticaria and anaphylaxis induced by cephalosporins have often been reported ( 7 ) . with regard to piperacillin
, there is only one published report of occupational asthma , rhinitis , and urticaria in a pharmaceutical worker ( 1 ) .
however , to our knowledge , there has been no report of occupational anaphylaxis induced by piperacillin .
furthermore , no evidence of a serum ige antibody response to piperacillin has been published .
this is thus the first report of occupational anaphylaxis due to piperacillin and detection of a serum ige response to a piperacillin - hsa conjugate .
the subject was a 24-yr - old female nurse who worked in the intensive care unit at a university hospital .
eighteen months later ( october 2004 ) , she experienced generalized urticaria with onset immediately and repeatedly after she had administered piperacillin
. nevertheless , she continued to work and finally experienced chest tightness , dizziness , sweating , generalized urticaria , abdominal pain , and diarrhea 10 min after giving a piperacillin injection in april 2005 .
her blood pressure was 80/50 mmhg with a pulse rate of 104 beats / min .
she had a previous history of atopic dermatitis , and her hand eczema had become exacerbated after she had begun to work at the hospital .
skin prick tests using 80 common inhalant and food allergens ( bencard , brentford , uk ) were all negative . we did not perform a skin prick test with piperacillin / tazobactam because of the risk of anaphylaxis ( 8) .
serum total ige , measured using an immuno - cap system ( phadia , uppsala , sweden ) , was 283 iu / ml . to assay piperacillin - specific serum ige and igg
, we prepared piperacillin - human serum albumin ( hsa ) conjugates in our laboratory and performed an elisa as described previously ( 11 ) .
piperacillin - specific ige and igg levels in an initial serum sample and the ige level in a serum sample obtained 5 yr later ( may 2010 ) were measured . in brief , the wells of a 96-well elisa microplate ( corning , new york , ny , usa ) were coated by incubation with piperacillin - hsa conjugate at 10 g / ml in phosphate - buffered saline ( pbs ) overnight at 4. after the wells were washed four times with 350 l of pbs containing 0.05% tween 20 ( pbs - t ; sigma - aldrich , st .
louis , mo , usa ) , nonspecific binding sites were blocked by incubation with 200 l of 10% fetal bovine serum ( fbs ; gibco / invitrogen , carlsbad , ca , usa ) in pbs at room temperature for 2 hr . after washing four times with 0.05% pbs - t , 50 l of diluted serum ( 1:3 ) were added to each well , followed by incubation for 2 hr at 37. the plate was washed four times with 0.05% pbs - t , then 100 l of goat anti - human ige antibody ( kirkegaard & perry laboratories , inc . ,
gaithersburg , md , usa ) diluted 1:1,000 with 10% fbs - pbs were added to each well , and the plate was incubated for 1 hr at room temperature .
after washing four times with 0.05% pbs - t , 100 l of alkaline phosphate - conjugated rabbit anti - goat igg antibody ( reserveaptm ; kirkegaard & perry laboratories ) diluted 1:500 with 10% fbs - pbs were added to each well , and the plate was incubated for 1 hr at room temperature .
pnpp ( p - nitro - phenyl phosphate ; sigma - aldrich ) substrate was added , and the reaction was stopped by the addition of 1 n naoh .
the optical density of the solution in each well was measured at 405 nm using a microplate reader ( synergy ht ; bio - tek , winooski , vt , usa ) .
the final absorbance value of piperacillin was determined by subtracting the absorbance value of a control crosslinker - hsa conjugate , which was obtained using a procedure identical to that described above , except that a crosslinker - hsa conjugate was used in place of piperacillin - hsa conjugate .
the positive cutoff value was determined as the mean plus three standard deviations of the absorbance values in serum samples from 11 unexposed , non - atopic healthy controls . for the inhibition elisa , increasing amounts ( 1 - 100 g / ml ) of free piperacillin , piperacillin - hsa conjugate , and hsa alone , dissolved in 10% fbs - pbs , were incubated with serum overnight , and an elisa was performed as described above .
the percentage inhibition was calculated as follows : % inhibition = ( 1 - absorbance with inhibitor / absorbance without inhibitor )  100 . for the elisa of piperacillin - specific serum igg levels , alkaline phosphate - conjugated rabbit anti - human igg antibody ( kirkegaard & perry laboratories )
was used instead of goat anti - human ige antibody and alkaline phosphate - conjugated rabbit anti - goat igg antibody .
a high level of specific ige antibody to piperacillin - hsa conjugate was detected in the initial serum sample from the subject ; however , this had decreased markedly after 5 yr ( fig .
1 ) . significant inhibition by both free piperacillin and piperacillin - hsa conjugate occurred in a dose - dependent manner , with minimal inhibition by hsa alone ( fig .
high levels of piperacillin - hsa conjugate - specific igg were also detected in the initial serum sample ( data not shown ) .
based on these findings , the subject was diagnosed with occupational anaphylaxis and urticaria caused by piperacillin .
the subject was transferred to a different department and has not again experienced anaphylaxis or urticaria .
skin prick tests using 80 common inhalant and food allergens ( bencard , brentford , uk ) were all negative . we did not perform a skin prick test with piperacillin / tazobactam because of the risk of anaphylaxis ( 8) .
serum total ige , measured using an immuno - cap system ( phadia , uppsala , sweden ) , was 283 iu / ml .
to assay piperacillin - specific serum ige and igg , we prepared piperacillin - human serum albumin ( hsa ) conjugates in our laboratory and performed an elisa as described previously ( 11 ) .
piperacillin - specific ige and igg levels in an initial serum sample and the ige level in a serum sample obtained 5 yr later ( may 2010 ) were measured . in brief , the wells of a 96-well elisa microplate ( corning , new york , ny , usa ) were coated by incubation with piperacillin - hsa conjugate at 10 g / ml in phosphate - buffered saline ( pbs ) overnight at 4. after the wells were washed four times with 350 l of pbs containing 0.05% tween 20 ( pbs - t ; sigma - aldrich , st .
louis , mo , usa ) , nonspecific binding sites were blocked by incubation with 200 l of 10% fetal bovine serum ( fbs ; gibco / invitrogen , carlsbad , ca , usa ) in pbs at room temperature for 2 hr . after washing four times with 0.05% pbs - t , 50 l of diluted serum ( 1:3 ) were added to each well , followed by incubation for 2 hr at 37. the plate was washed four times with 0.05% pbs - t , then 100 l of goat anti - human ige antibody ( kirkegaard & perry laboratories , inc . ,
gaithersburg , md , usa ) diluted 1:1,000 with 10% fbs - pbs were added to each well , and the plate was incubated for 1 hr at room temperature .
after washing four times with 0.05% pbs - t , 100 l of alkaline phosphate - conjugated rabbit anti - goat igg antibody ( reserveaptm ; kirkegaard & perry laboratories ) diluted 1:500 with 10% fbs - pbs were added to each well , and the plate was incubated for 1 hr at room temperature .
pnpp ( p - nitro - phenyl phosphate ; sigma - aldrich ) substrate was added , and the reaction was stopped by the addition of 1 n naoh .
the optical density of the solution in each well was measured at 405 nm using a microplate reader ( synergy ht ; bio - tek , winooski , vt , usa ) .
the final absorbance value of piperacillin was determined by subtracting the absorbance value of a control crosslinker - hsa conjugate , which was obtained using a procedure identical to that described above , except that a crosslinker - hsa conjugate was used in place of piperacillin - hsa conjugate .
the positive cutoff value was determined as the mean plus three standard deviations of the absorbance values in serum samples from 11 unexposed , non - atopic healthy controls . for the inhibition elisa , increasing amounts ( 1 - 100 g / ml ) of free piperacillin , piperacillin - hsa conjugate , and hsa alone , dissolved in 10% fbs - pbs , were incubated with serum overnight , and an elisa was performed as described above .
the percentage inhibition was calculated as follows : % inhibition = ( 1 - absorbance with inhibitor / absorbance without inhibitor )  100 . for the elisa of piperacillin - specific serum igg levels , alkaline phosphate - conjugated rabbit anti - human igg antibody ( kirkegaard & perry laboratories )
was used instead of goat anti - human ige antibody and alkaline phosphate - conjugated rabbit anti - goat igg antibody .
a high level of specific ige antibody to piperacillin - hsa conjugate was detected in the initial serum sample from the subject ; however , this had decreased markedly after 5 yr ( fig .
1 ) . significant inhibition by both free piperacillin and piperacillin - hsa conjugate occurred in a dose - dependent manner , with minimal inhibition by hsa alone ( fig .
high levels of piperacillin - hsa conjugate - specific igg were also detected in the initial serum sample ( data not shown ) .
based on these findings , the subject was diagnosed with occupational anaphylaxis and urticaria caused by piperacillin .
the subject was transferred to a different department and has not again experienced anaphylaxis or urticaria .
although we did not perform provocation and skin prick tests with piperacillin , we diagnosed this patient with occupational anaphylaxis because : 1 ) the temporal correlation between handling piperacillin and anaphylaxis ; 2 ) a history of recurrent urticaria after handling piperacillin ; and 3 ) detection of piperacillin - specific ige in the serum .
the complete disappearance of anaphylaxis and the reduction of serum ige after avoidance of piperacillin further support this diagnosis .
these findings suggest that occupational exposure to piperacillin induced ige - mediated anaphylaxis in this subject .
piperacillin can induce allergic reactions such as bronchial asthma , rhinitis ( 1 ) , urticaria , anaphylaxis ( 9 , 12 ) , immune hemolytic anemia ( 10 ) , maculopapular exanthem ( 13 , 14 ) , acute generalized exanthematous pulstulosis ( 15 ) , and drug hypersensitivity syndrome ( 16 ) .
several reports used skin prick , provocation , and serologic tests to demonstrate that piperacillin induced these reactions ( 1 , 9 , 10 ) .
a previous study reported that two of 311 tertiary hospital nurses showed a positive skin prick test to piperacillin ( 0.6% ) ( 17 ) .
two studies attempted to measure piperacillin - specific ige levels in serum ( 1 , 9 ) .
one involved a case of anaphylaxis , urticarial , and angioedema after intravenous piperacillin injection in an acute bronchitis patient , and the other was a case of occupational asthma , rhinitis , and urticaria induced by piperacillin powder exposure in a pharmaceutical worker . in those reports , a radioisotope test was used to detect ige levels , but the assay failed . in the present study , we successfully detected piperacillin - hsa conjugate - specific serum ige by elisa in a patient with suspected piperacillin - induced occupational anaphylaxis . furthermore , the binding specificity of the ige was confirmed by inhibition elisa . as significant inhibition by both free piperacillin and piperacillin - hsa conjugate
was detected , it is likely that the hapten component of piperacillin is the antigenic determinant .
high serum piperacillin - hsa conjugate - specific igg levels were also detected in this patient , while levels in controls were minimal ( data not shown ) .
although one study has sought to evaluate penicillin - specific igg levels in the sera of patients with penicillin allergies , the significance of igg levels remains unclear ( 18 ) .
further studies are needed to investigate the role of specific igg or its subclasses in occupational allergic diseases induced by piperacillin .
nontherapeutic exposure to antibiotics , including penicillins , can occur by various routes ( 3 ) .
contact with spilled drugs and inhalation of powder or foam are the most common routes .
exposure by the contact route usually induces contact urticaria ( 4 ) , anaphylaxis ( 19 ) , or dermatitis ( 5 ) , whereas the inhalation route induces most commonly asthma or rhinitis ( 2 , 20 ) .
piperacillin , an intravenous antibiotic , is provided as a powder in a bottle and should be dissolved prior to administration . when dissolved , piperacillin develops little foam , but emits a more obvious odor than other intravenous antibiotics .
the subject in the present report sometimes suffered from urticaria without direct contact with the solution , suggesting that inhalation of evaporated piperacillin may be the major route of sensitization .
however , considering the reported exacerbation of hand eczema that occurred prior to the anaphylaxis , we speculate that exposure by direct contact with piperacillin solution may also have been a factor . in conclusion , we report a case of a nurse with anaphylaxis and generalized urticaria caused by piperacillin and mediated by an interaction of ige with the hapten of piperacillin .
sensitization was suspected to have occurred through inhalation of piperacillin vapor . given that an elisa was capable of detecting piperacillin - specific serum ige , this system could be used for detecting piperacillin - hsa conjugate - specific ige in the serum of occupational anaphylaxis or asymptomatic sensitized patients .
 this is the first reported detection of serum ige antibody to piperacillin - human serum albumin ( hsa ) conjugate in a patient presenting with anaphylaxis that developed after occupational exposure . a 24-yr - old nurse , who had worked at a university hospital for 2 yr , experienced chest tightness , dizziness , generalized urticaria , abdominal pain , and diarrhea 10 min after administering a piperacillin injection . 
 she had previously suffered from atopic dermatitis . 
 a skin prick test for common inhalant allergens was entirely negative ; in contrast , her serum total ige was elevated ( 283 iu / ml ) 
 . a high level of piperacillin - specific serum ige was detected by elisa using piperacillin - hsa conjugate . 
 significant inhibition upon addition of both free piperacillin and piperacillin - hsa conjugate was detected by inhibition elisa . 
 these data suggest that piperacillin exposure in the workplace can induce occupational anaphylaxis and urticaria mediated by an interaction of ige with the hapten of piperacillin . 
INTRODUCTION
CASE DESCRIPTION
Skin prick tests and measurement of serum total IgE
Assay of serum IgE and IgG levels
DISCUSSION