Purification of brush border membrane by thiocyanate treatment

Abstract

Rat small intestinal brush border membranes are purified from brush borders by homogenization in relatively high concentrations of thiocyanate salts (0.56 m LiSCN, 0.41 m NaSCN, or 0.52 m KSCN), removal of this salt, and differential centrifugation to separate cytoskeletal material from membranes. The marker enzyme, sucrase, is enriched 98-fold in the final membranes over the starting homogenate of intestinal scrapings at a yield of about 20%. The isolated membranes are capable of secondary active sodium-dependent glucose transport as demonstrated by sodium gradient-supported overshooting glucose uptake.