Saturday, January 7, 2017

TOMAHAQ -- Rapid, targeted, triggered, multiplexed quan with attomol sensitivity!!


YEAAAAHHHH!!!!!!  I'm so psyched!!!  Before I get ahead of myself, what I'm about to ramble about is this paper in press at Molecular Cell from Erickson and Rose et al.,



What is it? Remember these papers? 


Just as a reminder, these guys hacked a Q Exactive and spiked in heavy peptide internal standards for the peptides they were interested in. When the QE sees the heavy standards then it starts doing high sensitivity PRMs in the mass range where the peptides they're interested in should be. They've done follow-ups and the method works like you'd think -- far more targets monitored than you can with an unaltered QE and with superb beyond QQQ level sensitivity.

TOMAHAQ takes this idea several steps further.

1) You don't have to make heavy peptide standards!  You make a mix of the peptides you are interested in and TMT-0 label them. Then you take your normal TMT-10 plex labeled samples and run them with this method.

TMT-0 has a mass of 224.something. TMT10 has a mass of 229.something. When the Tribrid sees the synthetic peptide it just has to move up 5 m/z and collect ions for MS/MS.

2) Unlike the Gallien et al., method -- this is TMT! You don't have to collect 10 scans across the peak to get your quan. SO...you can use a lot of fill time if you want -- and get unheard of levels of sensitivity!

If I have one criticism of this amazing new method -- it is that they could have pushed this SO much further. Hey -- I get it, it is a brilliant brand new methodology -- get it out there! But they did multiple fragmentations (8 I think!) of their targets. With TMT SPS3, this is very expensive from a cycle time sense and is going to limit the amount of sensitivity or number of targets. I'm gonna guess that this group is running this technique at much higher levels in their lab right now.

3) It is TMT!!! Every run is getting them super sensitive targeted quantification of 10 samples!!!  How good is this methodology? They targeted 69 proteins across the ENTIRE NCI-60 cell line set (60 different cancer cell lines) in TRIPLICATE in 48 hours!!

If you did this with a triple quad, with 20 minute runs (counting loading and equilibration and all that) this would take 60 hours. And...whole cell lysate SRMs in 20 minutes...look like crap (sorry, but they do.)

4) Also, unlike the Gallien et al., method -- YOU CAN DO THIS ONE!!  (If you have a Fusion or Lumos, that is...) you don't need to rewrite the operating software for the instrument. You just set it up in the normal Fusion software. I think you can do it in 2.0, but you can definitely set it up in 2.1!

EDIT 1/16/17: TOMAHAQ can only be set up in Fusion software 2.1.

How much do I like this method? I'm gonna risk my street cred and look like a casual NBA fan by ending this post with the generic go to for best tomahawk dunk ever, with Vinsanity himself!  That's how much I like this paper....makes sense as a scoring method in my head.... (I really like this method!)


2 comments:

  1. See Thermo Scientific video
    Multiplexing Targeted Proteomics Assays: Triggered by Offset Multiplexed Accurate mass High resolution Absolute Quantification (TOMAHAQ) on a Lumos Tribrid

    http://analyteguru.com/videos/multiplexing-targeted-proteomics-assays-triggered-by-offset-multiplexed-accurate-mass-high-resolution-absolute-quantification-tomahaq-on-a-lumos-tribrid/

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  2. See Analyteguru video
    A Strategy to Combine Sample Multiplexing with Targeted Proteomics Assays for High-Throughput Protein Signature Characterization

    http://analyteguru.com/videos/multiplexing-targeted-proteomics-assays-triggered-by-offset-multiplexed-accurate-mass-high-resolution-absolute-quantification-tomahaq-on-a-lumos-tribrid/

    ReplyDelete