Problems with densitometry!....for WESTERN BLOTS

[Jeff Rawlings]

In article <gjmadlam-2203951154280001 at 97.233.med.umich.edu>,
gjmadlam at umich.edu (Gerard ) 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'm not sure if the chemiluminescent systems are linear (they might be - I
just don't know). The solution I used was to use I-125 labeled
secondaries, and develop the blots in a phosphorimager. Both of those
steps should be linear and quantitative.

Cheers,
--Jeff

-- 
Jeff Rawlings                                   Internet Consultant
rawlings at silcom.com                                             and
rawlings at cco.caltech.edu                         Research Biologist