ISOLATION AND CHARACTERIZATION OF PLASMA-MEMBRANES FROM TRANSPLANTABLE HUMAN ASTROCYTOMA, OAT-CELL CARCINOMA, AND MELANOMAS

  • 1 January 1981
    • journal article
    • research article
    • Vol. 41  (10) , 4031-4038
Abstract
Purified plasma membranes were obtained from 5 transplantable human tumors: a Grade IV astrocytoma, an oat cell carcinoma and 3 melanomas. Plasma membrane fractions were isolated from tumor homogenates by differential and discontinuous sucrose gradient centrifugation. Determination of enzyme activities indicated that the plasma membranes were enriched 10-20-fold with respect to 5''-nucleotidase, NAD glycohydrolase, Mg2+-activated nucleoside triphosphatase and sialic acid. Specific activities of nearly all the enzymes varied with the individual tumors, even among tumors of the same type, i.e., the melanomas. EM of the plasma membrane fractions showed smooth single-membrane vesicles with slight contamination by lysosomes. These membranes are suitable for comparative biochemical studies and for the preparation of tumor-specific monoclonal antibodies. Plasma membranes from all 5 tumors contained very high Mg2+-ATPase activities. The Na+-K+-ATPase was a minor component of the total ATPase of these membranes (< 30%). The major component was an ATPase exhibiting similar activity toward several nucleoside triphosphates. The activity of such a nucleoside triphosphatase has been correlated with tumorigenicity in cultured liver epithelial cells. The nucleoside triphosphatase of the plasma membranes of astrocytoma and oat cell carcinoma was stimulated from 50-100% by concanavalin A; ATPase of the melanoma plasma membranes was not or was only slightly stimulated. The different responses to concanavalin A could be due to differences in the ATPase molecules of the individual tumors or to the different environment of the ATPase.