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Background: Immediate allergic reactions to beta-lactam antibiotics are mediated by specific IgE antibodies. The Phadia CAP System FEIA is a commercial method for quantification of specific IgE. We wished to determine anti-beta-lactam IgE antibodies in patients without penicillin allergy but with high levels of total IgE., Methods: Sera from 41 patients (31 with high total IgE, 10 with low total IgE) were analyzed for IgE antibodies specific to penicilloyl G, penicilloyl V, amoxicilloyl and ampicilloyl using the CAP FEIA((R)) method that was available up to 2006. Seven sera that tested positive were rechecked in a new improved system available after 2006., Results: In patients without a history of penicillin allergy, the specificities of commercial tests for anti-beta-lactam IgE antibodies were 100%, 60%, 27% and 20% at total IgE levels of 8-263 kU/l, 500-664 kU/l, 1000-2000 kU/l and > 2000 kU/l, respectively. In seven retested sera, only 2 (28%) were still positive for penicillin-specific IgE antibody., Conclusion: Before 2006, tests for anti-beta-lactam IgE antibody in patients with total IgE > 500 kU/l were probably often false positive. Patients who were diagnosed as penicillin allergic before 2006 solely on the basis of a positive CAP FEIA test for specific IgE should be considered for diagnostic reevaluation.

Background: Penicillin administration is usually contraindicated in penicillin-allergic patients with positive skin test results., Objective: To examine whether penicillin oral challenge for patients with a history of remote non-life-threatening allergic reaction to penicillin can be well tolerated irrespective of skin test results., Methods: In a prospective open-label trial, 8,702 individuals were screened between November 1998 and January 2000. Of 687 patients with a non-life-threatening allergic reaction to penicillin, occurring longer than 3 years earlier, 169 were enrolled. Regardless of the response to penicillin skin testing, patients received the usual 1-day dosage of penicillin and amoxicillin, on 2 separate occasions. Two to 6 years later, a follow-up was conducted to assess the outcomes of further penicillin administration., Results: A total of 272 combined skin tests and oral challenges were performed on 169 patients. Among 137 challenges with a positive skin test result and 135 patients with a negative skin test result, 9 (6.6%) and 5 (3.7%) (P = .29), respectively, developed a mild rash to oral challenge. At follow-up, 2 to 6 years afterward, 3 of 55 patients (5.5%) who were given a full treatment course of penicillin developed a mild skin eruption., Conclusions: Positive penicillin skin test results for patients with a remote history of non-life-threatening allergic reaction to penicillin were not associated with a greater prevalence of adverse reactions to oral challenge with penicillin than negative results. Because skin testing is considered the gold standard and the safest method for predicting tolerance to penicillin administration, oral penicillin challenge may be used as a diagnostic method only in these specific patients when skin testing is not feasible.

In this study we have explored the presence of cross reactivity to penicillin and ampicillin in patients with immediate allergic reactions caused by amoxicillin. Skin test results with amoxicillin were positive in 66.66% and in the remaining 33.34% were negative. In 81.48% patients we observed positive results of skin tests with penicillin and/or ampicillin. Only in 14.48% of the patients we observed selective response to amoxicillin. On the basis of obtained results we conclude that sensitivity of skin tests with amoxicillin is rather moderate and confirmation of amoxicillin hypersensitivity can be obtained using skin tests with other penicillins. In our material selective amoxicillin hypersensitivity can be defined as a relatively rare phenomenon. These data should have the important implications in antibiotics' selection in patients with amoxicillin allergy.

Background: Literature reports dealing with cell-mediated allergy to betalactams have appeared with increasing frequency in the last years., Objective: To evaluate patients with such reactions and to identify cross-reactivities among betalactams in order to provide safe guidelines for their further clinical management., Methods: Thirty consecutive subjects with cell-mediated allergy to betalactams (history of adverse reactions to these antibiotics; serum total IgE within the normal range; absence of serum specific IgE antibodies to penicillin G and V, amoxicillin, and ampicillin; negative skin tests with a wide pattern of betalactam preparations; and positive patch-test to at least one betalactam antigenic determinant) were investigated. The subjects admitted to the study were patch tested with a wide variety of betalactam preparations in order to identify alternative molecules tolerated by the patient. To better evaluate the cross-reactivity pattern, tolerance challenges with patch-negative betalactams were also performed in each subject., Results: Both specific IgE and skin tests were negative in all patients. The skin biopsies performed on the positive patch-tested area in four patients showed a clear T-lymphocyte, CD4+-type infiltrate, thus definitely proving the occurrence of a cell-mediated response. A total of 44 adverse reactions (mean: 1.47 episodes for each patient) were reported in history, with a mean interval of 15 hours after betalactam administration. The reported symptoms were mainly cutaneous (maculo-papular rash and urticaria) and the responsible drugs were chiefly aminopenicillins (86.4% of cases) and penicillin G (9.1%). We were able to identify three separate groups of patients on the basis of clinical history, patch-test, and tolerance challenge pattern: allergy to the side chain of aminopenicillins in 16 patients (53.3%); allergy to the thiazolidine ring in 3 patients (10.0%); undetermined specificity in the remainder 11 patients (36.7%). Cross-reactivity among different betalactam molecules (revealed by positive tolerance tests performed with patch-negative betalactams) was found in 4.8% of cases only (23.3% of all investigated patients). This fact demonstrates a very high (95.2%) predictive value of a negative patch-test in excluding the occurrence of a cross-reactivity. The mis-match between patch and tolerance tests was observed in 3 out of 178 cases only (1.7% of cases, 10.5% of patients) in groups A and B, and in as much as 12.2% of cases (45.5% of subjects) in group C (P < .05)., Conclusions: Delayed allergy to betalactams (mainly to aminopenicillins) may be exerted by a cell-mediated response. Patch tests and tolerance challenges are extremely useful and safe for diagnosis and further clinical treatment of these patients, helping to identify safe alternative betalactam molecules that could be successfully tolerated by the allergic subjects.

Background: Ampicillin (AMP) is a drug that has been prescribed extensively. Reactions that have been reported include exanthema, desquamative contact eczema, urticaria and anaphylaxis. Experimental evidence indicates that the side chain of AMP is a structure that may induce a selective immune response either at the humoral or lymphocyte T-cell level. With regard to IgE reactions, the selectivity and specificity of the response needs to be studied in humans., Objectives: To study the specificity of the IgE response in a group of subjects who had an immediate allergic reaction after the administration of AMP., Methods: Subjects developing an immediate response (anaphylaxis or urticaria) after the administration of AMP or an aminopenicillin derivative with the same side chain as AMP were studied. Skin tests were made to determinants generated from benzyl penicillin (BP): benzyl penicilloyl (BPO) and minor determinant mixture (MDM), as well as amoxicillin (AX) and AMP. Specific IgE antibodies were determined to benzyl penicilloyl polylisine (BPO-PLL), amoxicilloyl-polylisine (AX-PLL) and ampicilloyl-polylisine (AMP-PLL). The specificity of the IgE antibody response was studied by RAST and RAST inhibition. Subjects were classified in three categories: group A: those who were skin test and/or RAST positive to determinants derived from benzylpenicillin, group B: those who were negative to determinants derived from benzylpenicillin but were skin test and/or RAST positive to determinants derived from AX and AMP and group C: those who were exclusively positive to determinants derived from AMP., Results: A total of 48 subjects was included in the study. In group A there were 35 cases, in group B 10 cases, and in group C three cases. RAST inhibition studies showed that in some instances the side chain of AMP could induce specific responses with a variable degree of crossreactivity between BP and AX., Conclusions: Although AMP can induce an immediate IgE response in subjects allergic to betalactams and the structure of the side chain may contribute to the specificity of the response, our results indicate that in most instances crossreactivity with the other penicillins exists and that in the groups studied selective reactions to just AMP derived determinants were uncommon.

The purpose of this study was to determine if the sustained release of ampicillin from a biodegradable drug-delivery system (microencapsulated ampicillin anhydrate (MEAA)) will increase or decrease the intensity of a hypersensitivity reaction compared with that observed with free drug. Ovalbumin, which is known to elicit a marked hypersensitivity reaction in guinea pigs, and microencapsulated ovalbumin (MOVA) were tested in parallel with ampicillin and MEAA. Guinea pigs were sensitized biweekly by subcutaneous and intramuscular injections of ampicillin, MEAA, ovalbumin, MOVA or placebo microspheres (test articles), each mixed with Freund's adjuvant, and challenged 2 weeks later, intradermally, with the free compounds. In a separate set of experiments, guinea pigs were sensitized by implantation of the same agents in the caudal thigh of anaesthetized animals. Skin allergic reactions were tested at 1 and 3 weeks following local implantation of the test articles. Sera of sensitized guinea pigs were tested for specific IgG antibodies by enzyme-linked immunosorbent assay, and skin samples from the site of the inflammatory reaction were fixed, stained and evaluated histologically. Guinea pigs sensitized systemically with MEAA or MOVA showed smaller, but not statistically different skin allergic response than animals given corresponding free compounds. However, guinea pigs sensitized by local implantation of MEAA showed a significantly lower inflammatory response (P < 0.0001) than those given an equivalent dose of the free drug. Guinea pigs sensitized with placebo microspheres showed a low inflammatory skin reaction which was similar to those sensitized with all doses of MEAA. There was no significant difference in specific IgG antibody response in the sera of guinea pigs sensitized locally with either free or microencapsulated ampicillin or ovalbumin. Histology of skin revealed a milder inflammatory reaction with MEAA or MOVA than with ampicillin or ovalbumin, respectively. We conclude that the encapsulated ampicillin or ovalbumin and subsequent release of each agent will elicit a reduced hypersensitivity reaction in guinea pigs than will the free agent.

Pharmacological or immunological activation of human basophil induces the expression of membrane markers such as CD63 and the release of histamine and leukotriene C4 (LTC4). LTC4 release test has the advantages of a cellular test which is obviously closer to the physio-pathological reaction than the direct determination of circulating antibodies. Moreover, the determination of a newly formed mediator eliminates the spontaneous release of mediator by simple cellular damage. We present here, after having controlled the reliability of the method for aero-allergens, preliminary results of LTC4 release obtained for haptens (ampicillin, methylprednisolone and acetylsalicylic acid) which are in favour of the use of this method in the complex field of drug allergy diagnosis.

Ampicillin and benzylpenicillin conjugated to human serum albumin were used as immunogens in order to obtain antihaptenic IgG responses in outbred guinea pigs according to different schedules, all involving complete Freund's adjuvant. The individual responses were characterized by ELISA and by ELISA inhibition using ampicillin, benzylpenicillin, and carbenicillin peptidic conjugates for coating and for inhibition. In several instances, drastically reduced cross-reactivity and even its absence were observed, although the penicillin antigens differ only in the side-chain. The notion that the invariantly present thiazolidine ring will always provide significant binding to antibodies against all penicillins differing only in the side-chain has to be dropped. The experiments were performed in relation to newer findings of clinical penicillin-allergy skin testing which suggest that benzylpenicillin-based reagents alone are not able to detect or predict all reactions against semisynthetic penicillins. The experimental evidence here obtained corroborates this conclusion.

A patient with ampicillin-induced toxic epidermal necrolysis (TEN) showed pronounced delayed hypersensitivity to ampicillin sodium by both intradermal and patch tests. In addition, a positive lymphocyte transformation test to ampicillin was demonstrated by an in vitro study. On the basis of these findings and the data found in the literature, we believe that delayed hypersensitivity plays a crucial role in the development of drug-induced TEN. We recommend patch testing with suspected medication in such cases.

The dose-response relationship between the frequencies or concentration of exposure to powdered drug allergens and drug induced allergic onsets was studied in a pharmaceutical plant for 15 years from 1974 to 1984. The subjects were 41 male workers and the target allergens were two kinds of anti-inflammatory enzymes (Bromelain and Trypsin) and three kinds of antibiotics (Ampicillin, Amoxycillin and Cephalexin). The allergic onsets were confirmed by periodic allergological examinations and occasional clinical findings. Statistical analysis was made by the person-year method. The results showed that in workers who had any allergic history, the incidence rates of allergic onsets increased with elevation in the frequencies or concentration of exposure to these allergens, while in the cases without such history, the incidence rates increased only in those with high frequencies of exposure to the allergens. The findings suggest that the incidence rates of occupational drug allergy were dependent on the frequencies and concentration of exposure to allergens.

A case presentation describes a young woman with a history of two reactions to ampicillin therapy and a reproducible skin test reaction of intermediate timing that disappeared within 48 hours. The skin test response was to ampicillin only and not to other penicillin-related skin test reagents. Tests for serum IgE and IgG antibody to ampicillin were negative. The histology was that of a mononuclear and neutrophilic cellular infiltrate with neutrophil margination in the vessels. There was no immunoglobulin, complement, or fibrin deposition. The skin test reaction began and ended earlier than would be expected for a classic delayed hypersensitivity reaction. It is considered to be an isolated late cutaneous response but cannot yet be designated a late cutaneous allergic response. Reactants characteristic of an Arthus reaction were not present, and no alternative immunologic basis was confirmed.

35 individuals showing reactions to penicillin of anaphylactic shock, angioedema or urticaria were investigated. Their skin sensitivity was analysed using 16 different penicillin derivatives. In addition, the content of circulating reagins against the penicilloyl structure in the patient's sera were analysed using RAST. 17 of the patients had negative skin reactions and RAST results to all substances tested. The other 18 were skin test-positive to at least one derivative but showed markedly heterogeneous patterns of skin reactivity. 14 had positive reactions against penicilloyl structures accompanied by anti-penicilloyl reagins. Four patients showed doubtful reactions only to penicillin or penicilloate and/or penilloate. These patients also had very low levels of reagins against penicilloyl in their sera. Positive skin test results using monovalent penicillin derivatives such as penicillin, penicilloate, penilloate, penicilloyl amide, penicilloyl-formyl-lysine, penicillamine, which cannot form a multivalent antigen with penicillyol specificity, indicated formation of other derivatives of importance in penicillin allergy, e.g., penicillamine protein conjugates. Three patients showed skin reactions to ampicillin polymer and two to benzyl-penicillin polymer. The skin tests performed with the penicillin derivatives used do not seem to give more information on the sensitivity of the patients than does the RAST using penicilloyl structures.

Antibody responses to penicilloyl were recorded in infants and children treated for about 1 week with intravenous ampicillin. In this single-blind, randomized study two commercially available preparations were compared, one of high-grade purity (less than 0.1 microgram antigen per gram) and the other slightly contaminated with high molecular weight proteinaceous material (1.5 microgram antigen per gram) according to a radioimmunoassay. The results showed no significant increase in the antibody titers in any of the patient groups. The immunogenic properties of high molecular weight proteinaceous impurities isolated from phenoxymethyl penicillin during manufacture were tested in rabbits given daily subcutaneous injections with 0.1-10 micrograms of the antigen over 10 day periods. When the 10-day period was repeated, the rabbits injected with more than 2 micrograms of antigen responded with both IgM/IgG and IgE antibodies. The clinical as well as the animal study indicates that antigen impurities in the preparations of the order of 2 micrograms per gram or less do not elicit a significant antibody response. However, the study in rabbits demonstrates that high molecular weight impurities can induce penicillin allergy if present in about 10-fold higher quantities than those usually found in the commercial penicillin preparations of today.

The present communication reveals a relationship between the epitope density of penicilloylated protein antigens and their antigenic activities in a radioimmunoassay (RIA), in passive cutaneous anaphylaxis (PCA) and in inducing antibody formation in mice. In the RIA and PCA a critical number of 2-4 penicilloyl residues per protein molecule was noted. At this level small changes in the number of substituents considerably influenced the antigenic activities. The molecular weight and the nature of the carrier proteins, myoglobin, bovine serum albumin (BSA) and dimeric BSA also affected the threshold concentration for efficient antigenic activity. The results with the RIA and PCA were significantly correlated to each other. Using penicilloylated BSA as immunizing antigen in mice it was found that an epitope number higher than 11 penicilloyl residues per protein molecule induced significant antibody formation after a single injection. Antigens with a lower degree of penicilloyl substitution were less immunogenic. An antigen carrying 0.6 penicilloyl residues per BSA molecule did not induce penicilloyl-specific antibodies even after three injections. The capacity of heavily penicilloylated proteins to induce and elicit penicillin allergy as revealed by the present results stresses the importance of limiting their presence in penicillin preparations.

Using a radioimmunoassay, it was shown that commercially available ampicillin preparations often contain penicilloylated high molecular weight impurities. These possess immunological activities and stimulate penicilloyl-specific antibody formation in mice treated according to a therapeutic schedule. Using purified and experimentally contaminated preparations it was also found that exposure of the animals to Escherichia coli and Bordetella pertussis bacteria could increase the antibody formation to small amounts of impurities. In addition, penicilloylated antigen could be recorded in serum from treated animals. The antigen formed by penicilloylation in vivo, however, was very weak and did not induce much antibody formation when injected together with Freund's adjuvant in mice or rabbits.

The effects of the different acyl side chains of azidocillin, ampicillin and benzylpenicillin on the immunogenic potency of penicilloylated antigens as well as on the specificity of the developed antibodies were investigated in CBA mice. The antigens used were penicilloylated bovine gammaglobulin (BGG), bovine serum albumin (BSA) and human serum albumin (HSA). Immunization was performed with injection of high doses of antigens together with adjuvant or by injection of minute amounts of antigen over periods of 10 days. IgE antibodies were recorded with PCA in rats and IgG antibodies were measured with a double antibody assay. The nature of the carrier as well as the number of epitopes was found to influence the development of antibodies irrespective of the immunization schedule used. The immunogenic activity of the PO-BSA antigens was related to the epitope density. The PO-BSA antigens were, in contrast to the PO-BGG antigens, weak immunogens in the CBA mice. The acyl side chains of the different penicillins influenced the induction and specificity of the IgE antibody responses obtained after daily treatment.

This communication presents a sensitive and discriminative method for detection of protein impurities in penicillin preparations. Antibodies against various penicilloyl derivatives of high avidities and specificities raised in rabbits were coupled to microcrystalline cellulose. The amount of penicilloyl antigen present in a sample was calculated from the relative uptake of a radioiodinated penicilloylated albumin competing with the sample for binding to the antipenicilloyl immunosorbent. As little as 0.048 pmol/ml of penicilloylated human serum albumin could be detected. The accuracy of the determination was within +/- 23% (p less than 0.05). The pronounced specificities against the penicillin side chains demonstrated by the various immunosorbents were not displayed by the antibodies in passive cutaneous anaphylaxis experiments in guinea pigs. Furthermore, the immunosorbents showed the same pattern of specificity against monomeric penicillins as for penicilloylated proteins, but the former were considerably less efficiently recorded. The relatively small quantities of protein impurities in penicillin preparations, however, necessitated a separation from penicillin prior to analyses with the RIA. This was accomplished by fractionation on Sephadex G-50 fine, ginving a recovery of 80-90% of 0.1-2.5 ppm of penicilloylated protein.

Immunization of Syrian female hamsters with benzylpenicillin and ampicillin resulted in positive skin reactions of the retarded type and decreased titers of the blood antihistamine factor (AHF) in a part of the posterity. Penicillin allergy was in particular observed in the posterity of the female hamsters immunited before the pregnancy. The state of allergy to penicillins was found in the posterity of the female hamsters with both the positive and negative skin reactions on immunization during the 2nd half of the pregnancy.

Nineteen well-characterized penicillin-allergic patients were investigated for their sensitivity to cephalosporins containing potentially cross-reactive side chains. All patients were administered cephamandole parenterally and, if this was tolerated, a course of oral cephaloridine was administered. Only two patients responded to the cephamandole; none of the remaining patients reacted to cephaloridine. Benzylpenicilloyl RAST-inhibition studies with sera from three subjects who had not reacted to the cephalosporins demonstrated that cephamandole linked to proteins was capable of recognizing benzylpenicilloyl-specific IgE antibody. It is concluded that consideration of side chain structures can help to predict possible cross-reactions between penicillins and cephalosporins, but carefully controlled challenge tests are advisable before penicillin-allergic patients are treated with cephalosporins. In relation to cross-reacting potential, in vitro experimental studies are difficult to interpret and may in some circumstances overestimate the risk.