Selection of an anti-CD20, single-chain antibody by phage ELISA on fixed cells

Cloning the correct genes that code for antibody-variable domains from hybr idomas is often complicated by the presence of several immunoglobulin trans cripts, some of them arising from a myeloma cell line. For the rapid functi onal evaluation of recombinant antibody fragments against cell-surface anti gens, we established an efficient expression and screening system using pha gemid antibodies and fixed cells. VL and V-H-polymerase chain reaction (PCR ) products, amplified from hybridoma cDNA, were cloned into the phagemid ve ctor pSEX81. After transduction into E. coli and phage rescue, clones were tested for antigen binding using a phage-enzyme-linked immunosorbent assay (ELISA) procedure with whole cells fixed to ELISA wells. This procedure fac ilitated the successful cloning of a functional anti-CD20, single-chain ant ibody from hybridoma cDNA. The CD20 B-lymphocyte surface antigen expressed by B-cell lymphomas is an attractive target for cancer treatment using immu noconjugates or bi-specific antibodies.