PURIFICATION OF ANTIBODIES USING PROTEIN-L-BINDING FRAMEWORK STRUCTURES IN THE LIGHT-CHAIN VARIABLE DOMAIN

Protein L from the bacterial species Peptostreptoccus magnus binds spe cifically to the variable domain of Ig light chains, without interferi ng with the antigen-binding site. In this work a genetically engineere d fragment of protein L, including four of the repeated Ig-binding rep eat units, was employed for the purification of Ig from various source s. Thus, IgG, IgM, and IgA were purified from human and mouse serum in a single step using protein L-Sepharose affinity chromatography. More over, human and mouse monoclonal IgG, IgM, and IgA, and human IgG Fab fragments, as well as a mouse/human chimeric recombinant antibody, cou ld be purified from cultures of hybridoma cells or antibody-producing bacterial cells, with protein L-Sepharose. This was also the case with a humanized mouse antibody, in which mouse hypervariable antigen-bind ing regions had been introduced into a protein L-binding kappa subtype III human IgG. These experiments demonstrate that it is possible to e ngineer antibodies and antibody fragments (Fab, Fv) with protein L-bin ding framework regions, which can then be utilized in a protein L-base d purification protocol.