Moving on

Today is bitter sweet.  One of the best scientists I've ever worked with is leaving my team today to pursue bigger and better things.  While her skills will be greatly missed and extremely difficult to replace, I'm excited by the fact that we'll probably get to hang out more often since people generally don't need two Field Apps Scientists in one place and I tend to spend quite a bit of time in Boston.  And probably more now, since she won't be there to take most of the requests in the area.

Good luck with this next stage, Dr. Cardasis!!!!  And don't think you're going to get out of giving me lessons on instrument physics any time soon!

SparseQuant

Classical thinking says to do peptide quan right with a mass spec, you need a labeled peptide for each and every peptide that you are doing quan on.  The problem is that this can be crazy expensive.

Label free quan doesn't use require any fancy expensive labels, but the quan is relative and can be difficult to reproduce.

What we need is a compromise.  What if you combined the technology to make it expensive AND difficult to reproduce?  Just kidding!  Would that be a paper good enough for Nature Methods?  Probably, but this is even better.

This new paper in Nature Methods demonstrates the power of integrating just a few labeled peptides and using them for obtaining an improvement in your quan.  Cut the price, but still get better quan than you will see from label free. All the benefits of targeted quan with a whole lot less price!
             

Proteomics in personalized medicine

I just learned about these companies this week and I'm pretty excited about them.  These are both companies that are using proteomic biomarker assays to supplement personalized medicine.

Integrated diagnostics is based in Seattle and has a system called XPresys Lung that uses biomarkers to determine whether lung nodules are cancerous or benign.  Check it out!

OncoPlex DX is based in Rockville, MD and has tests for biomarkers for lung, breast, and colorectal cancers. Their website is here.

 I LOVE seeing proteomics moving toward the clinic.  So many good people are out there using global screens to find biomarkers and here are people using those biomarkers to benefit patients?

Proteomics WIN!!!

CompOmics

I ended up at a bar restaurant with some awesome scientists last night and one of the many things we ended up talking about was the number and quality of free resources that are popping up in proteomics.  Its getting to the point now where just about everything you'd want to do is out there.  The trick, a lot of the time, is in hunting down the software that will do what you want.  Unfortunately, the other trick is in getting it installed because sometimes our bioinformatics friends forget that not all of us know R and Perl, or use Linux.

Fortunately, this later problem seems to be getting better and I especially expect it to improve now that we have a precedent established of publications based on changing great freely available programs into easy to operature (and install) GUI programs.

To the topic at hand:  More free resources!  I was alerted to this one by a reader in India (thanks Dr. Sreelakshmi!!)

The CompOmics group in Ghent, Belgium has a great website with all sorts of free software as well as tutorials on how to use said software.  The software ranges from data parsers (from OMSSA or Mascot) to cell motility analyzers.  In short, tons of cool stuff!

Direct link to the site here!

Plos One new data access policy

As of March 1st, all Plos One journals require a new section at the end of the article you submit.  This is the data availability statement.  All authors must not only make all data available, but it must be specifically stated where this data is and how to obtain it.  For more information on the new Data Policy, click here.

ABRF 2014!

Woooooooo!!!!!!

N15 labeling in Proteome Discoverer

This question pops up a lot, despite the fact that this workflow is available on both the BRIMS portal and PlanetOrbitrap.  It never hurts to put it up in an additional place, right?

Question:  Can I do N15 labeling quan in Proteome Discoverer
Answer:  Yup!  But its a little tricky.

This link will take you to a Powerpoint with some instructions and an XML file that has a method I made for someone for PD 1.3.

Ultimately, the real trick is adding the modifications you need for each amino acid.  You need specify these mods uniquely by how many nitrogens are in each of your amino acids.

First start by updating your Unimod database.

Then add these labels.

Once these are in there, you can basically build the method based any of the precursor based quan methods (like SILAC). Comparing your light to your heavy.

Now, it gets a whole lot trickier if you are looking for incomplete incorporation.  You begin to run outside the realm of what you can do with the SILAC node.  What you need to do at that point is run your N15 mods as dynamic and use the precursor ion area detector and some database work will be necessary.  It certainly isn't impossible, but it will make for a nice long day of analysis.

The XML method in this file assumes that you are trying to compare two things, channel 1 with no N15 and channel two with 100% incorporation of N15 into each amino acid.  If you alkylate your cysteines you will need to make a custom modification that includes the mass shift of your N15 on cysteine as well as the mass shift of your alkylation (cause we can't yet do 2 mods/amino acid).

EDIT: Here is the new download link for the PD 1.3 slide deck.

More evidence -- DMSO study on the Q Exactive!

Early this morning I received an awesome Powerpoint describing results of another DMSO comparison.  This time it was for a TMT experiment performed on a Q Exactive instrument.

The work is by Dr. Henrik Johansson and Dr. Rui Branca of the Karolinska Institutet and my colleague Dr. Michaela Scigelova.

Guess what?  DMSO additives in buffer reproducibly increase peptide/proteins in the QE during nanospray applications.  Interestingly, it isn't to the same degree as the increases we have seen in the hybrid platforms.  I obtained permission to distribute the document this afternoon, so I'll put a direct link here until it is published on Planet Orbitrap.

Highlights:  
Adding DMSO led to 69% increase in +2 charge state peptides.  This is particularly cool, because TMT labeling tends to increase the peptide charge state (which can translate to a decrease in sequencing effiicency)

Adding DMSO led to a 17% increase in unique protein IDs.

Interestingly, comparison of the same sample ran twice, DMSO vs. normal buffer = new peptides!

Makes me wonder what those of you with super fancy multiple pump nanoLCs could do with this information....

Now, as always with this buffer additive -- I want to strongly caution you.  We do not have longitudinal data yet to fully realize the implications of this (or any other) buffer additive on nanoLC pumps, seals, or even the MS system itself.  Please follow the guidelines and specifications for your particular LC, source and instrument.  When in doubt, always please contact your respective vendors and technical support lines.  Also, please refer to my Disclaimer statement regarding this and any other advice you obtain through this blog.

You can download this incredibly informative and thorough analysis here.

Science fair...

Had to share this.  I'm guessing the parents aren't scientists....

Antibodypedia!

I was reading through JPR abstracts and saw the mention of this resource.  The article, however, wasn't open access, so they won't get any credit here.  (I don't want y'all thinking I read something that I really didn't, anyway.)

This resource is awesome and I wish it was around when I had to "validate" every proteomics result with some sort of antibody-based assay.

Here is how it works:

1) You enter your protein of interest and hit the search button

The antibodypedia comes back with some ontology information on your protein, a list of orthologs and how many antibodies are commercially available.

By clicking on the antibodies that are available, you get an expanded report leading you right to said antibodies!

Not only do you get the provider AND the catalog number, but you get an easy reference table for what that antibody is useful for; Western, immunocytochemistry, precipitation, and on and on!

The solid green dot means that the antibodypedia has verified evidence that the antibody works for that type of assay.  The 3/4 dots means the manufacturer shows evidence that the antibody works for that assay, but it hasn't been individually confirmed.  The 1/4 dot means that the manufacturer says it will work, but there is no evidence out there.

How easy is that?!?!  I've been a little uninspired this week so far, but this is exactly the kind of stuff I want to write about.

You can directly link to the antibodypedia here!