Characteristics of plasma membrane isolated from a mouse T lymphoma line: comparison after nitrogen cavitation, shearing, detergent treatment, and microvesiculation.
Plasma membrane was isolated from the mouse T lymphoma cell line WEHI-22 using four different methods of cell disruption followed by centrifugal fractionation. Disruption by nitrogen cavitation or by shearing with a cell pump produced plasma membrane vesicles of similar buoyant density (1.10 g/ml) and morphological appearance. Few C-type virus particles were present. Cell disruption with 2% Tween-40 produced membrane vesicles of similar morphology but lower density (1.09 g/ml). All of the above preparations resulted in vesicles with aggregated intramembranous particles after freeze fracture. Microvesiculation with sublytic concentration of a lysophosphatidylcholine analog (ET-12-H) (0.0032% w/v) produced small membrane vesicles which could be isolated without differential centrifugation. However, these had a slightly higher density than vesicles prepared by cavitation or shearing and were contaminated by virus particles. Unlike the other preparations, vesicles prepared with ET-12-H had dispersed intramembranous particles. The enzyme gamma-glutamyl transferase was enriched from 20- to 45-fold in the membrane preparations and proved a suitable plasma membrane marker for these cells whose 5'-nucleotidase content is very low.
D000818:Animals; D002458:Cell Fractionation / Q000379:methods*; D002460:Cell Line; D002462:Cell Membrane / Q000032:analysis / Q000201:enzymology / Q000648:ultrastructure*; D002499:Centrifugation, Density Gradient; D008223:Lymphoma; D008244:Lysophosphatidylcholines; D008563:Membrane Lipids / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis; D051379:Mice; D011136:Polysorbates; D013601:T-Lymphocytes; D005723:gamma-Glutamyltransferase / Q000378:metabolism
D008244:Lysophosphatidylcholines; D008563:Membrane Lipids; D008565:Membrane Proteins; D011136:Polysorbates; D005723:gamma-Glutamyltransferase