Tuesday, April 7, 2015

Label free quan on a Q Exactive global proteomics sample!

Alright, this paper is really cool.  I know these days when people think global label free quan (both inside and outside the field) they think Data Independent Acquisition approaches like pSMART or SWATCH. Despite some existing evidence to the contrary, I think I still fall in the camp where I'd prefer to see a high resolution MS1 comparison between two different samples.  I see the power of DIA, obviously, but precursor ion intensity is a very valid quantification method when done correctly and it makes me when I see evidence of people doing it right.

A great example is this new paper from Tali Shalit et. al., that just came out in JPR.  In this study these researchers performed label free quantification on a super complex sample using a Q Exactive.  How complex?  They added stuff to a HeLa digest and separated it in 1 dimension!  Using the same amount of HeLa, they spiked in different concentrations of an E.coli digest.  This gave them a great way to test the accuracy of their quantification over the full dynamic range of the instrument with known values.

How good is the Q Exactive?  Incredible, of course.  <10% CVs across the entire dynamic range and measurements that compare accurately to the known values all the way into the limits of detection.

Now, one of the cool things analyzed was the importance of the processing software.  That data is there in the RAW file but not every piece of software is created equal.  In this analysis they looked at MaxQuant's iBAQ algorithm (I know, I probably didn't capitalize the right letters...I can hear my coffee machine violently extracting caffeine now...) and Expressionist.

Expressionist has come up in my conversations a few times recently. And the series of events go like this.  Someone says something about Expressionist.  I write it on my hand or EverNote or something.  Then  I see the word later and Google it.  And I come up with an entire page of pictures like the monstrosity above. Then I remember that I've done this repeatedly and that, whatever it is, it is something I've looked up incorrectly multiple times.  Adding the term "proteomics" to the toolbar gets me to the GeneData Expressionist page (here.)

I haven't had a chance to check this software out personally yet.  I do like the tutorial videos and I've watched a couple of them (hey, I don't tell you what to do with your free time!) Whenever someone speaks in an accent that isn't my native one, I assume they are smarter than me. The guy doing the videos sounds super smart so I just assume it is nice software.  But this is the first comparison I've seen with Expressionist vs another software package.

In the case of this experiment, Expressionist appears to be a whole lot better than the label free quantification options in MaxQuant.  This may be an artifact of the sheer complexity of this sample, but maybe not.  You'll have to dig through the Supplemental material yourself cause I've got other stuff to do today ;)

All around, this is a very nice paper and a great short coffee read!  Particularly if this is the kind of study that you want to make a focal point in your lab.

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