storing western blots

Paul N Hengen pnh at fcs260c2.ncifcrf.gov
Wed Mar 22 18:59:37 EST 1995


Gerard (gjmadlam at umich.edu) wrote:

| I've just made a strange discovery when developing by blots.  I found that
| when I expose my blots to film, after using ECL, the bands that I get for
| shorter exposures aren't always "lighter" than the bands I get for longer
| exposures.  They are sometimes darker!!!!  This also happens in reverse
| (of course).  I was just wondering if anyone else has had this problem
| and, if so, what solutions are there.  Thanks.

I had a similar experience with the Southern Lights [TM] chemiluminescent kit
from Tropix. Two gels run, blotted, and detected under identical conditions
with the same amount of biotinylated DNA loaded onto the gel required different
times of exposure to get similar intensity of bands. One took only 15 min.,
while the other took over an hour. I can't explain it. I know I did everything
exactly the same, and the DNA is end-labeled (no probing or hybridization).
Do you think this could be the same problem? I'm curious to know if you meant
film exposed to the same blot for shorter time was darker, or that different
blots take longer to get a similar intensity.

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* Paul N. Hengen, Ph.D.                           /--------------------------/*
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