Preparation and characterization of anti‐framework antibodies to the heavy chain variable region (V H ) of mouse immunoglobulins

Abstract The heavy chain variable portion of mouse myeloma protein MOPC‐315 (α, λ 2 ) was obtained from its Fv fragment (J. Hochman, D. Inbar and D. Givol, Biochemistry 1973. 12 : 1130.) and was used to immunize rabbits. Rabbit anti‐V H antibodies precipitated V H , Fv and the intact protein 315. The V H region specificity of these antibodies was evident from the line of identity in agar gel diffusion between Fv and M315, and from the lack of precipitation with another a chain of XRPC‐25 or with light (L) chains. Radioimmunoassay binding and inhibition studies demonstrated that anti‐V H 315 antibodies cross‐react with heavy (H) chains derived from various mouse myeloma proteins (MOPC‐460, XRPC‐25, MOPC‐104E, UPC‐10, TEPC‐15, HOPC‐8, MOPC‐167) as well as with H chains derived from mouse IgG. These H chains can inhibit 90–100% of the binding of V H 315 by anti‐V H indicating extensive sharing of antigenic determinants by various V H regions. The relative affinity of anti‐V H for various H chains was 20–1000‐fold weaker than its affinity for the MOPC‐315 H chain. The cross‐reaction of the various H chains was independent of antibody specificity, subclass, subgroup or C H allotype. This suggests that anti‐V H may be considered as anti‐V framework antibody. Anti‐V H 315 cross‐reacts with isolated H chains but not with intact molecules and can therefore be used to detect V H determinants only in the absence of chains.