Wednesday, August 1, 2018

What ratio of isobaric labeling reagent to peptide is critical?

(Shoutout to A.J.Bureta for assembling part of this image and posting it under the Creative Commons Share-A-Like 3 where "free to modify" is rule number 2 {Hopefully my modifications aren't too obvious}).

Okay -- so I've noticed a trend recently -- I first noticed it right before ASMS in this great new paper. While at ASMS I hunted down a couple of studies that used isobaric labeling technologies and spoke to them about the ratio of peptides to labeling reagent they are using. The final straw was last week when I got to talk to a somewhat famous researcher in isobaric labeling techniques who referred me to some earlier works like this one is the question. When I pull up a protocol for isobaric labeling reagents, I'll find a recommended ratio of 8x labeling reagent to every 1x of peptides. In a protocol for another I find 10x.

Do we need that much reagent? In the first paper above, they clearly use a 2x labeling reagent to 1x peptides. In conversations with other researchers it sounds like 2x or 4x is becoming more of the secret (hidden deep in the methods section) norm. I can imagine that 8x probably does label closer to 100% of the peptides -- but if you could get 95% labeled with 2x or 97% with 4x  or 99% labeled with 8x --- umm..... that last 4%...ummmm --

[insert unrelated gif here]

....totally unrelated...

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