Immunoblotting a calcium-binding protein

Rick Davis rick at unity.ncsu.edu
Thu Aug 24 13:57:42 EST 1995


I am attempting to detect a plant calcium-binding protein (calmodulin?) on
a western blot with little success.  A bacterial recombinant protein from
the isolated gene does appear on a western, but the calcium-binding
protein extracted from plant tissue does not (concentration problem?). 
Silver-staining of the gel with the plant extract appears to indicate that
the desired protein is present, but difficulty in the transfer (alhtough
not present in gel after electroblot?) or binding to the membrane may be
the problem -- have tried nitrocellulose and PVDF membranes.  Has anyone
had similar difficulties in immunoblots of calcium-binding proteins -- any
suggestions (buffer conditions, different membranes, etc.)?

-- 
rick at unity.ncsu.edu



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