Friday, October 4, 2019

Maximizing Ion Trap MS/MS Acquisitions! (Get 12% more IDs out of any system?!?!)

I took a semester off from school in undergrad. Up to that point my priorities were running around in a costume (I was my school's mascot), dancing and tumbling poorly in said costume at basketball games I got into for free and had the best seat possible for (! no seat!) and going to a lot of dumb parties. I lost a scholarship and ran a gas line trimmer through one summer and the following semester before sobering up and swearing I'd never get anything but an A for the rest of my life.

Then I took Dr. Iulia Lazar's mass spectrometry class in grad school (Bioanalytical instrumentation) and got the first B since I made that oath (and last one of my life so far).

I bring this up because I'm an oversharer -- BUT holy cow -- that B is my excuse for never even thinking of distorting this very basic tenet of tandem mass spectrometry we've all been taking for granted for a decade or two as this group describes in this awesome paper. After reading the abstract you will probably subconsciously start working on your excuse.

You don't live in Madison? ...probably valid....

For the generation of newer people to our field who have been exclusively using high resolution things like an Orbitrap -- it doesn't matter what your mass range is there. All the ions get in and the frequency is read while they're in there. A 1.4 Da SIM scan at 35,000 resolution takes the same amount of time as a 35,000 resolution full scan.

Okay -- concept #1 -- so a scanning device like a quadrupole OR AN ION TRAP has a certain speed per mass range it covers. In an ion trap all the ions it can hold are there and then it sequentially ejects them. (In a quad it sequentially lets them through, but that doesn't matter here -- but if you try to do a full scan on a triple quad you'll be shocked by how slow it is). Ejection is super fast, but it isn't instantaneous. It takes longer to eject a full scan than it does a SIM scan (overheads and all sorts of other things do play a role because they're not instantaneous either, but let's keep going)

#2 -- there is some stuff in every MS/MS scan that we don't reeeeaaaaaaaly care about. Probably ions that are on the high and low end of the m/z. Do people in DIA typically let their scan windows go all the way to the bottom? Do you care about that arginine? Really? Probably not! DIA scan ranges are, as far as I can tell, always static.

#3 -- We're basically always letting the instrument determine on the fly what scan range to use. For the "high-low" stuff where you're doing, for example, Orbitrap MS1 followed by (or synchronous with, probably a combination of the two) MS/MS in an ion trap, the automatic settings are just taking your center point and the "1/3 (3/8?) cutoff rule" as the low mass.

What this group did was sit back and say -- wait a minute -- are we possibly hurting our results by letting an arbitrary rule of physics (that whole stability diagram thing) decide how we do our biological measurements?

YES. Yes we are. How does 12% strike you? A 12% gain in unique identifications!!! People will buy entirely new systems for a 12% increase in unique identifications.

Unless I'm missing something, this should be applicable to every ion trap system doing proteomics out there and should apply to anything where data dependent acquisition is used in complex mixtures.

If you're using an ion trap to identify anything complex you should definitely check this fantastic new study out.

1 comment:

  1. Would this make any difference using a quad instead of an ion trap?