Frequency of precursor cells against the enzyme β‐galactosidase An estimate of the BALB/c strain antibody repertoire

Abstract

A method has been developed for detecting anti-β-galactosidase antibodies after isoelectric focusing in thin layers of polyacrylamide gel. By “staining” with wild-type enzyme, all antibodies against β-galactosidase are detected, while a subset of antibodies able to activate a mutant enzyme is detected by staining with that enzyme. Limiting dilutions of β-galactosidase-primed or unprimed spleen cells of BALB/c mice were transferred together with antigen into sublethally irra-diated syngeneic hosts. The limiting role of the precursor B cells has been judged by the analysis of the clonal distribution of galactosidase-specific antibodies in recipient sera. The frequency of anti-wild-type β-galactosidase precursor cells was one in 0.42 × 10⁶ in the primed and one in 0.93 × 10⁶ in the unprimed spleen. The frequency of precursor cells for antibodies activating the mutant enzyme was one in 1.5 × 1O⁶ in the primed and one in 4.6 × 10⁶ in the unprimed spleen. Therefore four and five times less anti-B (mutant) than anti-B (wild-type) precursor cells exist in the spleens of primed and unprimed BALB/c mice, respectively. Comparing 51 clones derived from one primed donor mouse, it was possible to demonstrate that at least 43 different mutant-β-galactosidase-activating antibodies can be produced in one mouse. Comparing these 43 clones with 27 clones derived from another donor mouse, only one clone seemed to be common to both mice. From this the repertoire of the BALB/c strain has been estimated to consist of over 1000 different mutant enzyme-activating antibodies.