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western blot problem - Blank film from WB with ECL (Mar/31/2004 )

Did anybody experience such a problem as this? We did western blot, 1st and 2nd antibody at room temperature, then using ECL plus from Amersham. But we keep getting blank film after development, not every blot, but once a while.

We did troubleshooting and we know it is not due to bad film, bad antibody or bad ECL reagents. We suspect a certain reagent in our lab has problem, such as buffer. But it is still striking to see blank film. Did anybody can kindly give any suggestions?


What do you mean blank film? Do you there is no chemiluminascence signal at all or it appears all over the menbrane?



ECL films are two-sided, are'nt they? An emulsion side and an enhancer side. Wrong side against the signal source gives you blanks. Alternately, are your fixer and developer solutions getting interchanged by someone?