Parallel Reaction Monitoring -- Do a ton of SRMs at once!

I love SRM.  Hell, everyone loves SRMs.  Nothing beats getting your ion of interest and identifying it with 1) retention time 2) mass, and 3) fragment ion(s).  The problem is that they are sloooooowwww to set up because method optimization is such a drag. You've got to find the right fragment ion and collision energy to make it show up at the highest efficiency (if someone hasn't done it for you -- on your particular instrument, of course). It isn't such an issue in proteomics, but in small molecule work SRM collision energies for particular compounds can be the biggest secrets in the world.

That's why this paper in MCP lab from Amelia Pierson et al., of the  Koon lab is so cool!

 Parallel reaction monitoring uses the targeted multiplex MS2 method on the Q Exactive and essentially lets you get a bunch of SRMs (or MRMs, depending on what you're calling it) all at the same time.  Not only is it faster, but because the Q Exactive has an inclusion list of up to 5,000 ions all selectable by retention time, with the right chromatography system you can validate hundreds of observations in one go. MS based virtual western blots in realistic time may actually be a reality! 

There are some huge advantages here --
1) You are getting high resolution fragments!
2) You are getting a lot of them (all of them?) at once so you can be selective in processing (if you've got a co-eluting compound of striking similarity, you can ignore that fragment ion and focus on ones that are truly unique
3) Less up front setup time!

EDIT: Holy cow. This post was live for 7 years with a really awful typo in it. Thank you to any person who ever saw this and ever talked to me or took me serious about anything in regards to mass spectrometry ever again. Fixed....

Mass Spectrometer on Curiosity Part 3: Conversation with a small molecule expert

Yesterday, I had the chance to run my thoughts on the MS system on Curiosity by a small molecule triple quad expert.  His thoughts really changed my perspective on this.  Apparently +/- 1 Da isn't all that bad for a quadrupole instrument.  Yes, there are better ones out there (such as ones produced by my employer  that can do +/- 0.2 Da without big sensitivity losses), but most +/- 1 Da is pretty much the industry standard.  While most quads can narrow down to smaller mass windows, the sensitivity losses are so severe that we it is completely impractical.
So, while the MS device on Curiosity isn't cutting edge, it isn't as primitive as I originally thought.  Makes you wonder, though, why they just didn't buy one from Thermo...

Mass Spectrometer on Curiosity Part II, more 70s technology?

Curiosity is a historic achievement.  It is the biggest and most complex instrument we have ever landed on another planet.  Virtually every T-shirt I have purchased since it landed has had a NASA logo on it.  I am very very proud of the scientists and engineers who have made this possible.  I am, however, very disappointed in the level of sophistication of the Mass Spectrometer on the Rover.  

This is a schematic of SAM (sample analysis at Mars) lab.  The first thing you'll notice is that SAM uses a quadrupole mass spectrometer.  Sure.  This makes sense.  Quads are robust and can take a jarring without requiring constant recalibration. The mobile mass spectrometers the military uses for detecting chemical warfare agents are almost exclusively quad instruments.

The disappointment comes in when you look at the capabilities of this mass spectrometer. 

The above mass spectrum was taken from this paper detailing the calibration of the quadrupole unit from Curiosity.  I understand, we are taking this instrument and shooting it to another planet, but are we looking at mass accuracy within +/- 1 Da?  That is what I get.  1 Da?  Now the quad has a really interesting mass range, from 2 - 550 m/z.  That is a tough range to work with, but 1 Da?  A guy I've been working with this week has been working with small molecules <100 Da and has been consistently been pulling parts per billion mass accuracy. In my opinion, this is where our technology is today.  I'm sure there are reasons for this drop off, but I can't help but be a little disappointed.

Mass spectrometer on Curiosity part I, the disappointment...

If you've been following the NASA Curiosity project even a little, I'm sure you head about the recent statement and subsequent let down of the "history books" statement regarding Curiosity's recent discoveries. Prior to the let-down, I began to seriously wonder what real capabilities the mass spectrometer on the SAM (Sample Analysis at Mars) had for detecting life.  What I've found so far has been a little disappointing.

While Curiosity does carry a mass spectrometer, it doesn't appear to be a particularly advanced one, as the sensitivity of the instrument may not be any more than the instrument that arrived on the Viking 2, which landed on Mars in 1976, (3 years before I was born)!

There are advances, obviously, but according to the article from Jeff Bada in the link above, these advances are all in the sample preparation and ionization.  My guess is that these are advances to help combat the potential false-positive test that the Viking landers were so famous for, but are actually still a little controversial (link to be added when I find the paper...)

Part II will address the actual capabilities of the MS device and what it can/cannot detect.

Extending the mass range of an Orbitrap

I think we've all been hearing rumblings of this for months at all sorts of conferences, but the paper finally showed up in Nature Methods 2 weeks ago.  In this paper, Rose et. al., physically modify an Exactive plus instrument to extend the m/z mass range up to 12,000 Da.  The steps seems predictably complex to this Biologist, but for you engineering types it might not seem that bad.  For this study, I'm sure it didn't hurt that Dr. Makarov himself was one of the co-authors!

$150 for doing a mass spec survey?

Since I'm pretty sure I don't qualify (I do work for a vendor, after all), I can't really verify this, but it might be worth checking out.  This company, SurveyStat is offering $150 for users of mass spectrometers to participate in a survey.  Again, may be a scam -- I don't know.  But it might be worth checking out. $150!  Here is the link they sent me.  Let me know if it works!

MS Bioworks

Note to the MS Bioworks advertising company:  making the app was useful to you as well!

I am currently running through a lot of the new Apps available for proteomics/genomics that are available for the Apple and Droid devices.  Some are useless (see Smarty Mass Spectrometer), while others are pretty awesome (see Sparkplug!).  The MS Bioworks app falls in this second category.
What does it do?
1) In silico protein digestion -- look up proteins by the UniProt ID or by human gene symbol (10 enzymes to choose from, though no OmpT yet)
2) Peptide fragmentation -- really simple to use, several mods are available, b and y ions only
3) Nice converters for temperature, pressure, molarity and my favorite -- A TUBING VOLUME CALCULATOR!  Give it your capillary ID (in um or mm) and the length and it gives you your total volume.
4) BLASTp access via NCBI
5) Handy charts for exact masses of amino acids, modifications, etc.,

And who developed this really nice free App? I looked them up.
MS Bioworks is an MS services company in Ann Arbor with some extremely nice equipment and capabilities.  They can do everything from whole protein analysis to global PTMs to quantification via SILAC, iTRAQ/TMT, label free, and they are also set up for SRM/MRM.  Their lab is set up with triple quads, an Orbi Velos Pro and a Q Exactive.
Couple cutting edge equipment with the level of thoughtfulness they display in their little application and I think you have found a great lab for running your proteomics samples.  Check them out.  And if you do, let me know how it goes, I have a feeling you won't be disappointed.

Note:  The app appears to be available only for the apple devices.  I can't seem to find it for my Android tablet.

The Beer Proteome!

Now we're talking!  Beer proteomics!
A paper this month in the Journal of Proteomics by Gianluca Picariello, et. al., takes an in-depth look at the proteome of an Italian beer.  Apparently, this is a challenging prospect because the process of fermentation induces a number of modifications to the proteins present (including rampant deamidation).  In order to get around this and other obstacles, complex processing requirements were necessary:

On top of the proteomics, the researchers were interested in the effects of some of the proteins/peptides present in this beer on celiac's T cells.

Smarty Mass Spectrometer -- Don't buy.

If you are on the Apple or Google apps page and run across the Smarty Mass Spectrometer, keep on going and don't give them your hard earned money.  This app has a very vague description about a donated mass spectrometer, which leads you to believe that it has something to do with science or something.  It doesn't.  Either the programmers had no idea what a mass spectrum is, or they are just slapping words on vague apps to see what idiots (me...) will actually pay for them.  What it does is plot the color density of pictures that you upload into the app.  My Pug could write this dumb app.

Recently discovered PTM may be surprisingly common.

O-GlcNAc-6-phosphate is a recently discovered PTM that was previously only identified on a single protein found in rat neurons.  A paper this month in MCP from Hannes Hahne and Bernard Kuster demonstrates that this modification is a whole lot more common.  By tracing specific reporter ions produced by the degradation of the mod during CID, they were able to confirm the presence of this modification on a number of different mouse neuronal proteins.  They also confirm that this modification is found on some human proteins.