A cationic detergent, cetyltrimethylammonium bromide (CTAB), selectively dissociates the intermediate filament of the fibroblast.
We investigated conditions for selective solubilization of the intermediate filament (IF) of BHK-21 cells, and found that a cationic detergent, cetyltrimethylammonium bromide (CTAB), was effective for rapid dissociation of IF into the monomeric form. More selective dissociation was performed with a combination of CTAB and Tween 40. The CTAB-dissociated vimentins were unstable, but the breakdown of them was successfully blocked by leupeptin. Thus, with our extraction buffer, composed of 1% CTAB, 1% Tween 40, 10 mM Tris-HCl (pH 7.4) and 25 micrograms/ml leupeptin, almost all of the vimentin as well as the desmin were solubilized, while two thirds or more of actins were retained in the CTAB/Tween-insoluble fraction.
Maki
K
K
Faculty of Pharmaceutical Sciences, Kyoto University, Japan.
Sagara
J
J
Kawai
A
A
D000818:Animals; D002460:Cell Line; D000077286:Cetrimonium; D002593:Cetrimonium Compounds; D002850:Chromatography, Gel; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000648:ultrastructure; D015151:Immunoblotting; D007382:Intermediate Filaments / Q000648:ultrastructure*; D008970:Molecular Weight; D011480:Protease Inhibitors; D014746:Vimentin / Q000302:isolation & purification*
D002593:Cetrimonium Compounds; D003902:Detergents; D011480:Protease Inhibitors; D014746:Vimentin; D000077286:Cetrimonium