An improved plaque assay for mouse myeloma (MOPC 315) cells for use in studies of humoral and cell-mediated immunity.
Dinitrophenyl-bovine albumin was coupled at room temperature to sheep red blood cells in a procedure which minimized spontaneous lysis and allowed the preparation of large batches and their use for at least 3 weeks. The modified erythrocytes were used as a substrate for detecting local hemolytic plaques in agar by myeloma MOPC 315 cells, which secrete a paraprotein IgA with high affinity for dinitrophenyl ligand. Conditions maximizing the number of plaques formed by a given number of tumor cells were found to include coupling the erythrocytes at 1 mg/ml dinitrophenyl-bovine albumin with a molar ratio of about 50, and incubation with an amino-to-carboxy cross-linking agent, 1-ethyl-3(3 dimethyl aminopropyl) carbodiimide, at 2 mg/ml for 50 min. The method thus developed was employed to measure cellular and antibody-dependent immune reactions against the MOPC 315 cells. The experimental results show comparisons of the plaque technique with other measurements of tumor cell injury. The nature of the assay, which requires only 500 cells per plating, and which tests the synthetic capacity of single cells, suggests its use in experiments which limit the number of target cells, and in immune reactions causing injury, but not necessarily lysis, of the target cells.
D000818:Animals; D000912:Antibodies, Neoplasm; D002417:Cattle; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D004136:Dinitrobenzenes / Q000276:immunology; D004912:Erythrocytes / Q000276:immunology; D005022:Ethyldimethylaminopropyl Carbodiimide / Q000276:immunology; D006462:Hemolytic Plaque Technique; D007109:Immunity; D007111:Immunity, Cellular; D051379:Mice; D008807:Mice, Inbred BALB C; D009374:Neoplasms, Experimental / Q000276:immunology*; D012710:Serum Albumin, Bovine / Q000276:immunology; D012756:Sheep
D000912:Antibodies, Neoplasm; D004136:Dinitrobenzenes; D012710:Serum Albumin, Bovine; D005022:Ethyldimethylaminopropyl Carbodiimide