Simultaneous Immunoblotting Analysis with Activity Gel Electrophoresis and 2-D Gel Electrophoresis
Resource
Methods in Molecular Biology, 536, 23-33
Journal
Western Blotting(Methods in Molecular Biology)
Pages
61-72
Date Issued
2009
Date
2009
Author(s)
Kurien, Biji T.
Scofield, R. Hal
Abstract
Diffusion blotting method can couple immunoblotting analysis with another biochemical technique in a single polyacrylamide gel, however, with lower transfer efficiency as compared to the conventional electroblotting method. Thus, with diffusion blotting, protein blots can be obtained from an SDS polyacrylamide gel for zymography assay, from a native polyacrylamide gel for electrophoretic mobility shift assay (EMSA) or from a 2-D polyacrylamide gel for large-scale screening and identification of a protein marker. Thereafter, a particular signal in zymography, electrophoretic mobility shift assay, and 2-dimensional gel can be confirmed or identified by simultaneous immunoblotting analysis with a corresponding antiserum. These advantages make diffusion blotting desirable when partial loss of transfer efficiency can be tolerated or be compensated by a more sensitive immunodetection reaction using enhanced chemiluminescence detection.