Premium
Conditions for improved adsorption of calmodulin to nitrocellulose: Detection by 45 Ca binding
Author(s) -
Hincke Maxwell T.
Publication year - 1988
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150090704
Subject(s) - nitrocellulose , calmodulin , chemistry , adsorption , chromatography , sodium dodecyl sulfate , electroelution , detection limit , blot , calcium , potassium , biochemistry , membrane , enzyme , polyacrylamide gel electrophoresis , organic chemistry , gene
Abstract The 45 Ca filtration method which allows calcium‐binding proteins to be detected after their adsorption to nitrocellulose filters was used to test conditions for improved Western blotting of calmodulin. Our results indicate that dodecyl sulphate is not efficiently removed from calmodulin during electroelution in standard transfer buffers. This detergent disrupts binding of calmodulin to nitrocellulose. Electrotransfer in potassium buffer which precipitates dodecyl sulphate leads to a 6‐fold increase in calmoculin retention by nitrocellulose. With this and other modifications, the detection limit of the 45 Ca overlay method of Maruyama et al. (J. Biochem. 1984, 95 , 511–519) for calmodulin is increased to 0.2 μg per lane.