Just got a new LiCOR Odyssey Fc imaging system, which can do fluorescence multiplexing in near IR wavelengths. Total protein detection has become the normalization technique of choice (better than using a housekeeping protein like actin or GAPDH), and our sales rep told us about their in-house reversible stain. Revert, which can be used since it doesn't fluoresce in near IR. I believe it has a bluish color. My lab has some Pierce MemCode reversible stain that I've never used, but I think it stains a similar color. I'd like to use up what we have on hand before purchasing a new product. The components and steps sound similar and I'm wondering if the chemistry is the same. I want to know if anyone has had the opportunity to use MemCode and/or compare it to Revert? Thanks.
@Michael
Per your inquiry about Ponceau S, MemCode, and IR Western Blotting, here is a publication from an Odyssey customer at U Virginia, Charlottesville:
"An Analysis of Critical Factors for Quantitative Immunoblotting"
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4401487/
NOTE: This customer used a different type of Odyssey than the one you have. For technical information about REVERT, please contact your local LI-COR consultant.
Hope this helps.
Our lab manager developed a reversible total protein stain and wash that works with IR, and is MUCH cheaper than commercial solutions. Direct message me if you want the details. The solution is based on the following paper: Article Quantitation of protein on gels and blots by infrared fluore...
A new study in Molecular and Cellular Endocrinology compares the accuracy of housekeeping proteins to total protein staining with Revert for WB normalization. Might be of interest -Article Use of the REVERT ® total protein stain as a loading control...
I tried revert for the first time this week (it' is very expensive). you read it in the 700 channel, so doesnt that mean it does fluoresce in the IR spectrum? I also found it did not come off. ponceau at least comes off super quick in water, this stuff barely budged after 10 mins in NaOH and methanol (the max time allowed) I did see a lot come off in 5% BSA TBS blocker though (havent rescanned after blocker though to fully check, only after reverse solution. this is a visual observation based on the blue left on the membrane). i had 20ug loaded.
Licor provides a wash solution to take out the stain if you need to use the 700 channel for your protein of interest (and yes, it is in the IR spectrum). We also have a similar solution for our fastgreen variant of the total protein stain ( a protocol developed based on the following paper: Article Quantitation of protein on gels and blots by infrared fluore...
). In general, and without using the removal solution, we find that most of the stain washes out by the end of a standard protocol (block/ primary/secondary antibody application and washes between/after)
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