Purification of plasma membrane from Acanthamoeba castellanii.
A simple method for isolation of plasma membrane from Acanthamoeba using self-generating gradients of Percoll is described. To obtain a membrane marker, intact amoebae were radioiodinated and the distribution of the radiolabel was followed through the plasma membrane isolation procedure. The purity of isolated plasma membrane was assessed by enrichment of radiolabel, by electron microscopy, and by enzymatic assays for contaminating membranes. As judged from enrichment of radiolabel, a 37-fold purification of plasma membrane was obtained. We estimate that 80% of the total protein was from plasma membrane and 10% from membrane-associated actin.
Clarke
B J
BJ
Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, Bethesda, Maryland 20892.
Hohman
T C
TC
Bowers
B
B
D000048:Acanthamoeba / Q000648:ultrastructure*; D000199:Actins / Q000032:analysis; D000818:Animals; D002458:Cell Fractionation; D002462:Cell Membrane / Q000032:analysis / Q000648:ultrastructure; D002499:Centrifugation, Density Gradient; D008565:Membrane Proteins / Q000032:analysis; D008854:Microscopy, Electron
D000199:Actins; D008565:Membrane Proteins