GLYCOSYLATION ANALYSIS OF A POLYREACTIVE HUMAN MONOCLONAL IGG ANTIBODY DERIVED FROM A HUMAN-MOUSE HETEROHYBRIDOMA

Glycosylation of the human monoclonal IgG1 lambda antibody (mAb) CBGA1 was analysed by lectin blotting. The CBGA1 antibody binds to several antigens including donor self antigens, as detected by ELISA immunoblo tting techniques and an erythrocyte binding assay. The mAb producing c ell line was obtained by EBV transformation of peripheral blood lympho cytes of a healthy donor followed by fusion to the heteromyeloma cell line, CB-F7. The resulting heterohybridoma was cultivated in a hollow fibre bioreactor system. A bulk pool of 0.9 g antibody was produced. F ab and Fc fragments of the purified mAb were prepared and analysed. A noteworthy heterogeneity of CBGA1 and its fragments in SDS-PAGE and IE F was detected. We found glycosylation in the Fab fragment of CBGA1 in addition to the conserved glycosylation site in the Fc fragment at As n 297. Fab glycosylation was detected in both the Fd region and the la mbda-chain. The glycosylation pattern of the gamma-chain differs from that of the lambda-chain. Sequence analysis of the VH gene shows a pot ential N-glycosylation site located in framework III at position Asn 7 5.