Accession Number : ADA180670

Title :   Use of Fluorodinitrobenzene to Identify Monoclonal Antibodies Which are Suitable for Conjugation to Periodate-Oxidized Horseradish Peroxidase,

Corporate Author : WALTER REED ARMY INST OF RESEARCH WASHINGTON DC

Personal Author(s) : Burkot,T. R. ; Wirtz,R. A. ; Lyon,J.

Report Date : 1985

Pagination or Media Count : 7

Abstract : During the recent development of a double antibody enzyme-linked immunosorbent assay for detecting malaria sporozoites in mosquitoes it was found that a large number of potentially useful monoclonal antibodies lost their capacity for binding antigen after conjugation to periodate-oxidized horseradish peroxidase (HPO). Since HPO reacts with primary amino groups, we used a simple chemical reaction with fluorodinitrobenzene (FDNB) to determine if the loss of antigen binding was due to the requirement for unmodified primary amino groups in the binding site. FDNB-treated antibodies which reacted with antigen in an indirect fluorescent antibody assay (IFA) also yielded successful HPO-antibody conjugates. Conversely, those antibodies which did not react with antigen after treatment with FDNB failed to produce useful HPO-antibody conjugates. These data suggest that conjugation of oxidized HPO to primary amines in or near the antigen-combining site yields conjugates in which the antibody is inactive. Use of FDNB to predict the suitability of antibodies for conjugation to periodate-oxidized HPO may save a considerable amount of time over randomly selecting antibodies for conjugation and testing. (Reprints)

Descriptors :   *ANTIBODIES, SITES, ANTIGENS, LOSSES, CLONES, AMINES, REPRINTS, FLUORESCENT ANTIBODY TECHNIQUES, FLUORESCENT DYES, MALARIA, SPOROZOITES, CULICIDAE, CHEMICAL REACTIONS, IMMUNOASSAY, PEROXIDASES

Subject Categories : Biochemistry
      Medicine and Medical Research

Distribution Statement : APPROVED FOR PUBLIC RELEASE