Monoclonal antibody to the crystal induced chemotactic factor from human neutrophils detects similar immunoreactive and biologically active protein in promyelocytic HL-60 cells.
The crystal induced chemotactic factor (CCF) is a protein released by human neutrophils (PMN-CCF) that phagocytize monosodium urate or calcium pyrophosphate crystals. This protein has been thought to play an important role in crystal induced arthritis. In the present report, a mouse hybridoma cell line A2900C2.12 which specifically secretes monoclonal antibody (MAb) to CCF has been developed. The antibody secreted by the cell line is mostly IgG2b and positive for both the light chains kappa and lambda. The MAb has been purified by affinity chromatography using a protein-A column. The purified MAb was found to be over 95% pure based on the results from SDS-PAGE. The MAb has been used to demonstrate that a leukotactic protein is released from polymorphonuclear leukocytes (PMN) and promyelocytic HL-60 cells in response to a phagocytic or degranulating stimuli such as monosodium urate crystals or N-formyl-Met-Leu-Phe (FMLP), respectively. "Western blots" and silver nitrate staining of the purified leukotactic protein from PMNs and HL-60 cells show that these proteins have the same immunodeterminant site(s) and both appear to have a molecular weight (m.w.) of about 15,000. A simple purification protocol including immunoaffinity high pressure liquid chromatography (HPLC) has been described for purifying chemotactic proteins with the same antigenic determinant as the PMN-CCF.
Bhatt
A K
AK
Veterans Administration Medical Center, St. Louis, MO 63106.
Spilberg
I
I
D000818:Animals; D000911:Antibodies, Monoclonal; D002460:Cell Line; D002630:Chemotactic Factors / Q000276:immunology* / Q000302:isolation & purification; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D015473:Leukemia, Promyelocytic, Acute / Q000276:immunology; D051379:Mice; D009504:Neutrophils / Q000276:immunology*; D014407:Tumor Cells, Cultured / Q000276:immunology
D000911:Antibodies, Monoclonal; D002630:Chemotactic Factors