Establishment of a monoclonal antibody to plasma cells: a comparison with CD38 and PCA-1

Abstract

SUMMARYA new monoclonal antibody which recognizes plasma cells was developed by utilizing two myeloma cell lines, KMS12PE (12PE) and KMS12BM (12BM). eslublished from lhe pleural efTusion and bone marrow, respectively, of the same patient. Since I2BM expresses CD20, CD38, and PCA-I antigens, while 12PE has lost CD20. 12PE is considered to be phenotypically more mature than 12BM. The 12PE cells were used to immunize a BALB/c mouse and a MoAb was produced which was more reactive to 12PE thun to I2BM. Thus, a clone. D2, was obtained. On Western blotting, D2 detected a single band of 54 kD under both reduced and non-reduced conditions. This antigen was not detected by Western blotting in peripheral blood lymphocytes that had been stimulated with pokeweed mitogen (PWM) for 7 days or in those not so stimulated. On flow cytometry, D2 detected a myeloma cell line, RPMI 8226. Another myeloma cell line, U266, was negative for D2 antigen. Staining various cell lines by D2 and other antiplasma cell antibodies, PCA-1 and CD38, showed that D2 is distinct from PCA-1 and CD38. The fresh myeloma cells of 14 myeloma patients were stained by D2 and for other plasma cell antigens. D2 strongly stained three samples obtained from patients with clinically aggressive myeloma, while CD38 stained all cases e.xccpt one. PCA-1 was positive in nine samples and negative in five. PCA-1 expression was observed in plasma cells obtained from pleural effusion and peripheral blood, while PCA-1-negative cases were not found in such samples, suggesting that PCA-I expression was related to extramedullary invasion. The morphology of the myeloma eells, classified according to Greipp's criteria, showed that there was no correlation between plasma cell antigen expression and plasma cell morphology. Analysis of D2 antigen expression should provide more information about the heterogeneity of myeloma cells.

The identification of irreversible rituximab-resistant lymphoma caused by CD20 gene mutations

“…Retroviruses carrying wild type and deletion mutants of CD20 , together with mock construct, were produced with the transient retrovirus packaging cell line G3T-hi (Takara) according to the manufacturer's protocol. Packaged retrovirus vectors were then used to infect a myeloma cell line, KMS12PE, 4 with subsequent selection using 500 μg/ml G418.…”