I'm up bright and early (for Ben) this morning (let's not discuss details such as numbers on time keeping devices. It's early somewhere...) and MHCs are on my mind.
I haven't failed at identifying MHC peptides in around 6 years or so. I think the blame ended up falling on someone else, and to be honest, I didn't know what the heck I was looking at anyway. I knew that they weren't tryptic, they might completely lack basic residues and they might have PTMs all over them. I was treating them exactly like any other sample that would be that insanely miserable to work with.
Fast forward 6 years. MHCs are coming and I'm diving through the literature hoping to find THE MHC IDENTIFICATION WORKFLOW OF THE FUTURE that I dreamed of in the past.
And...well...hmm....HEY! Here is a recent review...
It covers what MHC peptides are and different ways people have
Honestly, when we were looking at this before I was convinced my biggest problem wasn't the mass spec. I was convinced it was the data analysis. This review has good news, by the way. My old lab is still doing MHC work and I think they are primarily doing this with PEAKS, which the review is solidly in favor of. I'll probably request a free trial to see how it does with the data we've acquired so far.
While I'm on the data processing topic, the great scientists in my facility have had success with the approach described in this recent study.
This is really elegant in it's simplicity. It takes the protein cleavage bias out of the data processing equation by allowing FASTAs to be generated that specifically contain just the MHC sequences. Sequest can't bias it's scoring toward the peptides ending in K or R when your FASTA entry doesn't contain any. Think of it like Xcomb for MCHs.
Okay -- and I made fun of the SWATH study mentioned in the review, but I'm actually printing that now as well. That study would require another work-around to the data processing problem -- spectral libraries! That could be a huge asset in searching MS/MS fragments from peptides that don't obey our super neat b/y ion rules!
I'm sorry if it seems like I made light of the complications of working with MHCs or studies where people have tried to advance this critical field in medicine. At least this sorry...
I'm glad to have a lot of stuff to read and to see how y'all have advanced the field and I'll be eagerly awaiting what is coming down the pipeline while we're struggling to work with these awful ions ourselves!
Wait...where are the UVPD MHC studies...?...someone is doing that, right?!?!? Someone living where there are cacti? (There are cacti in Austin, right? )