Monday, December 24, 2018

DIWA -- Dual Isolation Width Acquisition for Combating Ratio Suppression!

We're not done innovating, people! Smarter instrument methods keep coming in all the time, and DIWA is an awesome and elegant new example that you can read about here!

Also, an earlier pre-print version is available at biorXiV here. I'm not sure how I missed it. It would have been really helpful...stupid job getting in the way of me reading every paper that comes out, I guess....

Here is the idea -- if you are doing MS2 based isobaric labeling quan (TMT/iTRAQ) you have 2 choices:

1) A really narrow isolation window that will help you get less coeluting ions. (Obviously of varying efficacy depending on your complexity, actual m/z, etc.,)

2) A wider window that will allow you to catch your more of your peptide isotopes.

-- here is a cartoon I have for something we're working on --

If you use a 0.4Da isolation width on any isotope of these two coeluting peptide species, you're going to miss a TON of signal. Maybe the majority of it! Are you going to get enough signal to get an MS/MS that you can match a peptide out of? Maybe....?

Dual Isolation Width Acquisition (DIWA) says --

(In case you're wondering why you remember this -- we did the research -- this was a commercial for a grocery kit that contained both hard and soft shell tacos. --@ProteomicsNews -- not afraid to ask the hard questions)

Two MS2 scans were acquired for each ion selected for fragmentation on an Orbitrap Velos system. In order to sort the scans out, they had slightly different HCD energies. A wider isolation was used to get the best signal for identification and super narrow isolation was used for quan.

How'd it do? Impressively well.

You don't have to believe the authors.  You can get the files at PRIDE here when the full print of the paper goes live

Things to note -- this was TMT 6 plex. I'll always hold the great Orbi Velos I once had in high regard, but it's slooow for TMT 10/11-plex. 6-plex? No problem!

This can totally be applied to the other instruments, like the 2x faster Elite or Q Exactives! I'm not sure how to write it for the QEs, though.... I'll think on it and if I figure it out, I'll add it. (Definitely ping me if you have the method!)

There are other important points here such as the ability the narrow isolation spectra give you to do regression analysis to assess your isolation interference. And -- this sounds totally smart -- the authors mention the potential application of ion mobility as something they might be exploring?!?!

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