An isotopic antiglobulin test for quantitation of anti-sheep red blood cell antibodies of individual Ig classes in mice bearing different IgCH Haplotypes

Abstract

An isotopic antiglobulin test (IAT) for anti-sheep red blood cell (SRBC) antibodies of different immunoglobulin (Ig) classes was developed and used to measure levels of IgM, IgA, IgG 1, IgG 2a, IgG 2b and IgG 3 anti-SRBC antibody in sera from 12 inbred strains of mice. Characterization of the Ig class specific heteroantisera prepared for use in this assay revealed substantial amounts of antibodies reactive with idiotypic and allotypic determinants of IgG 1, IgG 2a, IgG 2b and IgA. Accordingly, class specific antibodies which do not discriminate among Ig molecules of different IgCH phenotypes were isolated from each heteroantiserum by absorption and elution from Ig's of selected strains. Levels of anti-SRBC antibody are expressed as units of antibody/ml relatvive to an anti-SRBC antiserrum reference standard. The use of conversion factors (estimates of the relative reactivity of each anti-Ig reagent) allows quantitative comparisons within and between IgG subclasses. This method allows measurements of nanogram amounts of specific anti-SRBC antibodies of a single Ig class, and simultaneous measurements of antibody levels in 6 Ig classes from a single 30 μl serum sample. Analysis of the responses of several strains of inbred mice indicates striking variations (5–31-fold) in the amounts and relative proportions of different Ig classes. The IgG subclass distributions observed appear to be associated with the IgCH but not the H-2 haplotypes of the strains studied.