Peroxidase-based chemiluminescence - best choice?
tguettou at newssun.med.miami.edu
Tue Mar 18 17:35:06 EST 1997
On Tue, 18 Mar 1997, Dima Klenchin wrote:
> Hello all,
> our lab is finally moving from I-125 to ECL-based Westerns.
> While I am sure every kit works, I am wondering what you guys
> would recommend to use most. Right now we can think of a reagents from
> Boehringer, Amersham, Pierce and NEN. Of course, other things
> being equal, the cheaper the better.
> Everyone who actually compared different "kits", please reply.
we compared Amersham ECL, Dupont Rennesaince, Pierce SuperSignal and
Pierce Super Signal Ultra.
Pierce SuperSignal won that comparance by far. It is the cheapest product
and has very good sensitivity and in my hands the lowest background.
You can expose for several hours. I like
the kit from Dupont still better than the one from Amersham, because it
is slightly more sensitive and also cheaper.
BTW there is no use in telling you detection limits in ug or ng of
protein, since they depend very much on the prim. and sec. antibody (in
case you use a sec. antobody). We compared them by using the same
conditions in our Western.
Finally, Pierce Super Signal Ultra is really the most sensitive
substrate. However, it's really expensive and I cannot nearly reach the
sensitivity they claim is possible (but see above!). As a comparance: 10
min exposure with Pierce Super Signal gives the same signal strength as
about 10-30s of exposure with the Ultra. In the case of the Ultra, I have
some trouble with the background. Exposures longer than 2 minutes blacken
my film completely. This might be due to the fact that I use the
secondary ab at 1:50000 and I probably should dilute it more.
Hope that helps.
( No affiliation with any of the companies)
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