Isolation and characterization of leukaemic B-lymphocytes: influence of anticoagulant on C3-receptor detection, humoral killing and capping of cell surface immunoglobulin.
Guinea pig L2C neoplastic B-lymphocytes processed from leukaemic blood collected into EDTA as anticoagulant exhibited high affinity rosetting for EAC mouse indicator cells. Partial blocking of the C3-receptors was apparent with blood collected into heparin or citrate as anticoagulant. This could be minimized by keeping the temperature at 4 degrees C and avoided by ensuring a high final concentration of citrate (greater than or equal to 40 mM). Results from experiments in vitro suggested that receptor blocking may have been a consequence of complement activation by the L2C cells occurring via the alternative pathway. No injury to the cells was evident with any of the anticoagulants used, and their susceptibility to antibody-dependent complement-mediated lysis was independent of anticoagulant and unaffected by the irregular presence of membrane-bound C3. In experiments where the antibody-induced redistribution of L2C surface immunoglobulin was investigated, it was found that cells processed from blood collected into EDTA showed poor ability to cap and interiorize their surface immunoglobulin in comparison with cells processed from heparinized or citrated blood. Inhibition of capping was found to be dependent on the concentration of EDTA and could be avoided by collecting blood into a large volume of low molarity EDTA as opposed to a small volume of high molarity EDTA. Some practical and theoretical consequences of our observations are discussed.
D000818:Animals; D000917:Antibody Formation; D000925:Anticoagulants; D001402:B-Lymphocytes / Q000276:immunology*; D001667:Binding, Competitive; D002417:Cattle; D002452:Cell Count; D002469:Cell Separation; D002471:Cell Transformation, Neoplastic; D003176:Complement C3; D004492:Edetic Acid; D004533:Egtazic Acid; D006168:Guinea Pigs; D007152:Immunologic Capping; D009374:Neoplasms, Experimental / Q000276:immunology; D011947:Receptors, Antigen, B-Cell; D011951:Receptors, Complement; D012397:Rosette Formation
D000925:Anticoagulants; D003176:Complement C3; D011947:Receptors, Antigen, B-Cell; D011951:Receptors, Complement; D004533:Egtazic Acid; D004492:Edetic Acid