AbstractThe polypeptide chain composition of class II antigens from LEW rat spleen cells was studied utilizing cross‐reactive mouse alloantiserum A. TH anti‐A.TL (specificity anti‐Iak) and the monoclonal antibodies MRC‐OX6 and MRC‐OX3 for immunoprecipitation. Two‐dimensional gel mapping of A. TH anti‐A. TL immunoprecipitates revealed that, as in the mouse, two groups of class II antigens exist corresponding to I‐A and I‐E locus equivalent structures. In the absence of reducing agents three monomeric chains α, 36 kDa (p36); γ, 33 kDa (p33); and β, 23 kDa (p23), were detected for I‐A equivalent antigens, whereas I‐E equivalent molecules separated into five monomeric chains: α, 37 kDa (p37); γ, 33 kDa (p33); and three β chains 28 (p28), 26 (p26) and 24 kDa (p24). One strong dimer component of disulfide‐linked γ chains was found to be associated with products of both loci. Although slightly different in molecular weight, γ chain corresponds to the nonpolymorphic murine invariant chain Ii.Both monoclonal antibodies recognized rat homologues of the murine I‐A products. Extensive sequential criss‐cross‐immunoprecipitation with subsequent 2‐dimensional O'Farrell analysis indicated that (a) MRC‐OX6 precipitated molecules which were not recognized by MRC‐OX3 andvice versa; (b) MRC‐OX6 precipitated a three‐polypeptide chain complex composed of the polypeptides p36, p33 and p23; (c) MRC‐OX3 precipitated a two‐chain complex composed of p36 and p23; and (d) the respective heavy (α) and light (β) chains of both complexes possess very similar physicochemical parameters, suggesting