Enzymatic solubilization of membrane immunoglobulin (M-Ig) from rabbit lymphocytes with phospholipase A2 (PL-A2) and phospholipase C (PL-C).
Enzymatic solubilization of M-Ig from rabbit lymph node cells was investigated using highly purified PL-A2 and PL-C. The combined treatment with PL-A2 (50 I.U./ml) and PL-C (20 I.U./ml) caused optimal solubilization of the membrane components from the 125I-labeled lymphocytes, but the treatment with either enzyme alone did not. The solubilized M-Ig was isolated and characterized as the membrane component which was specifically co-precipitable with homologous antigen-antibody complex. The solubilized M-Ig associated with some other membrane constituents was eluted in a void volume fraction by gelfiltration on a column of Sepharose 6B, and recovered at the interface between 30 and 40% sucrose layers by the density gradient centrifugation. The isolated component could be further separated by SDS-polyacrylamido gel electrophoresis into four radioactive polypeptides with apparent mol wt of 7 approximately 8 X 10(4), 4 approximately 5 X 10(4), 3 approximately 3.5 X 10(4) and 2 approximately 2.5 X 10(4), respectively. The results suggest that the enzymatic solubilization of lymphocyte M-Ig is a useful procedure to investigate further characteristics of M-Ig and their biological function related to the intracellular mechanisms of immune response.
D000818:Animals; D000936:Antigen-Antibody Complex; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D008214:Lymphocytes / Q000276:immunology*; D010740:Phospholipases / Q000378:metabolism*; D011817:Rabbits; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism*; D012995:Solubility; D013997:Time Factors
D000936:Antigen-Antibody Complex; D011947:Receptors, Antigen, B-Cell; D010740:Phospholipases