Comparison of the sensitivities of the Buehler test and the guinea pig maximization test for predictive testing of contact allergy.

International test guidelines, such as the Organisation for Economic Cooperation and Development (OECD) guideline #406, recommend 2 guinea pig methods for testing of the contact allergenic potential of chemicals: the Guinea Pig Maximization Test (GPMT) and the Buehler test. Previous comparisons between the methods suggested that the Buehler test was less sensitive than the GPMT although modified Buehler test protocols were used. Parallel GPMT and Buehler tests were conducted according to OECD guideline #406 using a multiple-dose design and test results were analysed using a standard logistic dose-response model. To compare the sensitivity of the 2 test procedures the test conditions were kept identical and the following chemicals with a range of sensitization potentials were tested: chloraniline, chlorhexidine, eugenol, formaldehyde, mercaptobenzothiazole and neomycin sulphate. Formaldehyde and neomycin sulphate were strong sensitizers in both tests. Mercaptobenzothiazole, eugenol and chloraniline were all strong sensitizers in the GPMT, eugenol and mercaptobenzothiazole were negative in the Buehler test and equivocal results were obtained with chloraniline. Chlorhexidine was negative in the GPMT and equivocal responses were obtained with the Buehler test. Higher induction concentrations were needed to show allergenicity in the Buehler test and for some allergens the Buehler test protocol was not sensitive enough to demonstrate allergenic potential.

Properties of a monoclonal antibody for the detection of abasic sites, a common DNA lesion.

The abasic site is one of the most frequent changes occurring in DNA and has been shown to be lethal and mutagenic. An abasic site in DNA can be tagged by reaction with O-4-nitrobenzylhydroxylamine (NBHA), resulting in the formation of an oxime linkage between the abasic site and the NBHA moiety. In order to measure NBHA-tagged abasic sites, a monoclonal antibody was elicited against a 5'-phosphodeoxyribosyl O-4-nitrobenzyl hydroxylamine-BSA conjugate. The antibody was specific for the NBHA residue as demonstrated by hapten inhibition, with IC50 values for 5'-phosphodeoxyribosyl-NBHA, deoxyribosyl-NBHA, ribosyl-NBHA and NBHA of 0.3 microM, 5 microM, 5 microM and 7 microM, respectively. Other haptens examined, including benzylhydroxylamine, 5'-phosphodeoxyribosyl-, deoxyribosyl-, and ribosyl-benzylhydroxylamine, showed no inhibition even at 1 mM. The antibody showed high specificity for NBHA-modified AP sites in DNA and exhibited no cross reactivity with normal DNA bases, otherwise-modified DNA bases or unmodified AP sites. Using a direct ELISA assay, the antibody detected 1 AP site (after NBHA-modification) per 10,000 base-pairs or approximately 10 femtomoles of AP sites in DNA. DNA lesions were detectable in 60Co gamma-irradiated DNA at a dose as low as 10 rad (0.1 Gy) and the production of antibody detectable sites was proportional to the gamma-ray dose. Since NBHA reacts with lesions containing an aldehyde group, the simplicity and sensitivity of the antibody assay should provide a useful method for the quantitation of AP sites or other DNA lesions containing an aldehyde group.

Diagnosis of sulfonamide hypersensitivity reactions by in-vitro "rechallenge" with hydroxylamine metabolites.

STUDY OBJECTIVE: To determine whether differences in in-vitro detoxification of sulfonamide-reactive metabolites can be detected among the lymphocytes from controls, patients with sulfonamide hypersensitivity reactions, and patients with nonhypersensitivity reactions to the sulfonamide agents. DESIGN: In-vitro toxicity assay on lymphocytes. SETTING: Clinics for adverse drug reactions in an adult and pediatric tertiary care center. PATIENTS: Peripheral blood lymphocytes were obtained from 46 normal volunteers and 76 patients referred to the clinic for assessment of adverse drug reactions to sulfonamide agents. Thirty-one patients had clinical histories consistent with a diagnosis of hypersensitivity reaction, whereas 45 patients had clinical histories felt to be inconsistent with a diagnosis of hypersensitivity reaction. INTERVENTIONS: Lymphocytes were assayed with tetrazolium to determine toxicity from the hydroxylamine of sulfamethoxazole. MEASUREMENTS AND MAIN RESULTS: The lymphocytes from patients with a history of hypersensitivity reactions showed markedly increased toxicity across a tenfold-concentration toxicity-concentration curve compared with those from controls and patients with a history of nonhypersensitivity reactions. These differences were highly significant (P less than 0.01). No difference was found between the toxicity shown by the lymphocytes from controls and that shown by the lymphocytes from patients with a history of nonhypersensitivity reactions. CONCLUSIONS: Metabolic differences in the production and detoxification of reactive metabolites of sulfonamide agents are important determinants of hypersensitivity reactions to these agents. These results suggest that the hydroxylamine derivative of sulfamethoxazole may be a reactive metabolite mediating these reactions. Sulfonamide hydroxylamines are useful in the diagnosis and study of the pathogenesis of hypersensitivity reactions to sulfonamide agents.

[Humoral and cellular immunity factors in peroral vaccination with an S. typhimurium antigenic complex]. / Gumoral'nye i kletochnye faktory immuniteta pri peroral'noi vaktsinatsii antigennym kompleksom S. typhimurium.

The role of individual immunity factors in protection from S. typhimurium infection was studied on mice immunized orally in a single administration with an antigenic complex obtained by the treatment of bacterial cells with hydroxylamine. The oral administration of this preparation induced the systemic and local transformation of the organism, which was manifested by the accumulation of antibody-producing cells in the immunocompetent organs (spleen, mesenteric lymph nodes) and the increase of the phagocytic activity of macrophages in peritoneal fluid. Cellular immunity factors were found to be important for the development of resistance to salmonellosis caused by S. typhimurium after oral immunization with the antigenic complex.