Protein blotting method for detection of nickel-binding proteins.
A technique is described for detection of nickel-binding proteins in tissue extracts by protein blotting and autoradiography. The proteins are separated by SDS-polyacrylamide gel electrophoresis and blotted onto a nitrocellulose membrane. The membrane is incubated with 63NiCl2 in a tris-HCl buffer that contains NaCl and CaCl2 to suppress non-specific Ni-binding. The membrane is washed to remove unbound 63Ni2+ and autoradiography is performed to visualize the 63Ni-binding proteins. The technique furnishes a sensitive means to detect 63Ni-binding proteins in tissue extracts, facilitating their isolation, characterization, and identification.
Lin
S M
SM
Department of Laboratory Medicine, University of Connecticut School of Medicine, Farmington 06032.
Hopfer
S M
SM
Brennan
S M
SM
Sunderman
F W
FW
D000818:Animals; D001345:Autoradiography; D015153:Blotting, Western; D002352:Carrier Proteins / Q000032:analysis*; D003600:Cytosol / Q000378:metabolism; D004563:Electrochemistry; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D066298:In Vitro Techniques; D008297:Male; D008970:Molecular Weight; D009532:Nickel / Q000378:metabolism*; D051381:Rats; D011916:Rats, Inbred F344; D013194:Staining and Labeling; D014981:Xenopus
D002352:Carrier Proteins; D009532:Nickel