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ECL Substrate Choices

ECL Chemiluminescence Western Blot

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2 replies to this topic

#1 Missle

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Posted 16 April 2014 - 09:02 AM

Hi.  I'm new to chemiluminescence western blots.  In searching for ECL substrates for use with HRP-conjugated secondary antibodies, I'm a little overwhelmed with the choices.  I'm pretty confident that I want a mid-sensitivity substrate to avoid the background issues with the ultra-sensitive substrates but there are so many choices.  Does anyone have favorites?

 

Thanks!



#2 bob1

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Posted 16 April 2014 - 12:25 PM

I use supersignal pico from Pierce.



#3 memari

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Posted 17 April 2014 - 09:49 AM

It depends on the amount of protein you have as a band.

We usualy start with

1- Least sensitive--->  Millipore Luminata Crescendo Western HRP substrate

http://www.millipore...LE/00112798.pdf

 

then, if there is no band of interest, we wash the membrane with T-TBS three times each 30 second. then we use this:

 

2- More sensitive---> Millipore Luminata Forte Western HRP substrate

http://www.millipore...LE/00112798.pdf

 

then, if there is no band of interest, we wash the membrane with T-TBS three times each 30 second. then we use this:

 

3- Most sensitive---> SuperSignal West Femto Chemiluminescent Substrate

http://www.piercenet...scent-substrate

 

but recently we found that CellSignal ECLs are very good and we are going to use these:

 

6883 SignalFire™ ECL Reagent

 

12630 SignalFire™ Plus ECL Reagent

 

12757 SignalFire™ Elite ECL Reagent

 

7003 20X LumiGLO® Reagent and 20X Peroxide


Edited by memari, 17 April 2014 - 09:54 AM.

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Babak Memari





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