Isolation of renal brush borders.
Methods are described to isolate intact brush borders and brush border membranes from renal cell homogenates. A rapid method yields sealed vesicles that reconstitute renal brush border transport. In one variation of this protocol, 10 to 20 mM CaCl2 or MgCl2 is added to aggregate non-brush border structures for subsequent removal by centrifugation. For analytical studies, guidance is provided for subsequent purification steps including preparative free-flow and aqueous two-phase partition. Marker enzymes and morphological parameters are included for assessment of yield and fraction purity.
Morré
D James
DJ
Purdue University, West Lafayette, Indiana.
Hammond
Timothy
T
Tulane University School of Medicine and Southeast Louisianna Veterans Health Care System, New Orleans, Louisianna.
D000818:Animals; D002021:Buffers; D002122:Calcium Chloride; D002458:Cell Fractionation / Q000379:methods*; D002499:Centrifugation, Density Gradient / Q000379:methods; D002501:Centrifugation, Zonal / Q000379:methods; D011232:Chemical Precipitation; D004396:Coloring Agents / Q000378:metabolism; D011992:Endosomes; D007672:Kidney Cortex / Q000648:ultrastructure*; D015636:Magnesium Chloride; D008297:Male; D008854:Microscopy, Electron / Q000379:methods; D008871:Microvilli / Q000201:enzymology; D011817:Rabbits; D051381:Rats; D017207:Rats, Sprague-Dawley; D012995:Solubility
D002021:Buffers; D004396:Coloring Agents; D015636:Magnesium Chloride; D002122:Calcium Chloride