Saved you a click, no real time targeting in this paper "iRT Assisted" paper

 

If you were thinking "cool, maybe someone made the great iRT triggering tools for Q Exactive available again!" maybe they have, but this isn't where they did it. 

This group did do targeted by standard PRM assay for the host-cell proteins that they are most worried about, and I've never seen this before. Generally host cell protein analysis is done by untargeted means - at least in the literature. The figures are nice, and they focus a lot of their reproducibility and ability to port these methods to other chromatography gradients, hypothetically. They also back up some of their results by ELISA and it's altogether not a bad study. 

How the reviewers let a helpful (not helpful! we have enough acronyms for methods! especially this one which is literally the node in the instrument interface) new acronym to get through to publication for someone following basically exactly verbatim the methods in the 2015 Skyline Tutorial for "how to do PRM", I do not know. Hopefully I can save you a click. 

Whoops. I forgot to put a direct link to the paper in. Too late now. 

Multi-instrument comparison of a proteomics process control pipeline!

 

How would your favorite mass spec do in a head to head in a process control pipeline? What's that? Something practical in manufacturing things, probably! Ask an AI, it will make some dumb shit up about it. Google's moron of an AI I can't seem to turn off just lied to me extensively about how many points different field goals in basketball count for. Unless I started the New Year with a stroke, I'm pretty sure I can't trust Doogle for anything now and I'm happy about this surprisingly inexpensive IOS migration in progress. 

What, was I typing about a paper? Yes! A preprint. This one! 

These authors prepared multiple combinations of two bacterium mixtures and used everyone's instruments. There's an Orbi, a TIMSTOF HT, a ZenoTOF, and even two Waters instruments. One of which I don't think I've ever even heard of. Either it's old or they called it something different in the UK, or I did have that stroke. Further evidence for the latter thing is that they somehow got Waters data into DIA-NN and MSStats! Except the Waters I don't know about, which was processed with Skyline. Sure, there are raw numbers here, but the cool part is that the authors focus on the pathways that they actually care about, and basically every instrument does really well in that regard, with the Vion being the clear outlier (on the bad side, but that might be Skyline 😜). 

Volcano enzymes for proteomics!

I kept forgetting this was a thing. Apparently, I even met some of the authors, but I was with the Director of Mass Spectrometry Innovation at some fancy place in California, and since we only hang out at conferences we tend to end up out until morning and I may not always form long-term memories at conferences with mass spec nerds everywhere. Which is funny when I pitch a collaboration and I'm all like "nice to meet you" and they are like "...ummm....what......" 

This is the original paper! 

This is the big picture! (Click to expand, or go to the paper link above)

AND the enzymes are surprisingly affordable! 

MASLD liver tissue AND plasma proteomics!

 

Thanks to our colleagues and affiliation with the Pittsburgh Liver Research Center we get access to a lot of human liver samples. Typically, however, this is because the liver is taken out of a person so that a new one can be put in. They don't typically take those livers out because they're in great shape. They're generally super extreme cases of bad livers. As you dig through the repositories you'll find that is often the case. Not a ton of helathy controls of livers or less terminal liver disease samples. 

Merry Xmas to me! 

Seven healthy controls WITH both liver homogenate and plasma proteomics to add to our hard drives? It is funny to me that the front page illustration shows a Sciex QQQ and the study was done with an Orbitrap Tribrid running DIA proteomics. I have one out or out soon where the other authors put an Agilent ICP-MS or something in their abstract graphic without consulting me on that change. 

All the data is up on MASSIVE here! 

What is really cool to me is that they only did the ones where they had matched plasma. Since the liver is a big organ with a tremendous interface with liquid blood stuff, the two should be closely related. And this group points out clear disparities between the two that are definitely worth my team thinking about. 

iFishMass - Direct (digested) nanoinfusion antibody (and ADC!) analysis!

 

Antibody and antibody drug conjugate (ADC) drugs are EVERYWHERE. You can't watch 3 minutes of YouTube or Television (is that still a thing? I can't believe YouTube still exists at all either, how hard would it be to replace it with something that was good?) 

Do you have to run 1 hour or 2 hour gradients of digests to work them out? If so, you sure couldn't keep up with a big multi-lot multi-batch generation facility.

Could you just digest and direct infuse? Probably! But how would you analyze those data? With iFishMass! 

They start off by doing some antibodies and ADCs with a standard nanoLC setup and then they move over to the NanoMate. Remember those? They are typically used for intact protein analysis. 

Turns out that on a Tribrid it works pretty great for simpler single protein digests. Some great news about the software for me is that you convert the data to a universal format before you put it into the software (one of the formats that starts with an X and ends in an ML. I forget which one. 

And iFishMass is up and available to run here. Funny point in the manuscript is where it says something like "the easiest way to install it is..." which is super handy and I'm glad the reviewers let them sneak in a helpful tip there. 

Joint pseudotime analysis of single cell transcript and proteomic data!

 

WHOA! Okay, I am just dropping this here so I can look at it later and see if it's helpful for us because I don't currently have a way to easily do this with protein data at all! (Translation: I haven't read this, but I'll see it and remember to later if it is here!) 

Spatial proteomics (with targeted technniques) at sub micron resolution! 30 proteins at a time!

 

I pitched mass spec based spatial proteomics a while back at a building where they have a whole ton of microscopes that are cooled with cryogens. Supposedly someone in the audience invented the whole idea of making a microscope super cold. A Wikipedia suggests that isn't all that unlikely. Right zipcode for sure. When they started asking questions about spatial resolution and I enthusiastically gave the best I'd ever seen, it sucked all the interest out of the entire room. 

For real, I think they considered not validating my parking. The microscopy people can't look at a lot of proteins at once, but when they look at them they want to be orders of magnitude below the best laser pulse our mass specs get.

What if you want to look at more than one protein at a time in microscopy? 

You get SUM-PAINT, I guess? They get some stupidly high spatial resolution while using oligonucleotide barcodes behind to label your targets. Ridiculously beautiful pictures and - I don't know if this is a great paper or approach, but it give some perspective on scale. There are a lot of 5nm pixels (their scale) inside of a 20 micrometer one....

Know a European Researcher who deserves an award? EuPA nominations open now!

 

5 categories! 5 prestigous awards from probably the coolest proteomics groups on the planet! 

You can nominate and find the rules and all the important stuff here! 

NanoDESI allows spatial intact protein complex analysis!?!

 

Well...this one looks like magic...

I was reading it on my phone yesterday, but I'm reasonably sure this was a custom Nano DESI source equipped on an Orbitrap Fusion 3 (Eclipse) or 4 (Assend). 

There are some ridiculously nice pictures in it, but if you're getting spatial localization of large proteins or medium sized protein complexes, there is some impressive mass spec wizardry going on here. Localizing a 185kDa protein in a human kidney??? Whoa. One of two really impressive spatial proteomics papers that dropped this weekend. The other one is in my wheelhouse a bit more and I'll probably get to my notes on it later. 

A spatial proteomic and phosphoproteomic map of liver mitochondria!

 

This isn't super new, but a colleague sent it to me and it's really really cool.

The liver is really weird and even though from a microscopic level it looks like a bag of square (they like "cuboidal") cells all stacked like bricks row after row forever, these cells are very different depending on where they are. These big bricks of cells are also packed full of mitochondria and may have hundreds of them per cell. This group used spatial sorting to get piles of hepatocytes from different zones THEN did mitochondrial enrichment THEN did (TMT) proteomics and phosphoproteomics.

There are big differences in mitochondria depending on where the cells are spatially in the liver. I was going through the methods and thought something like "wow! someone knew what they were doing! why don't I recognize any of these names?" I re-read the names. I know the 9th author. Wait. How is a mostly proteomics paper...PI...is....9th...author....meh...probably politics stuff..... There is pretty ....confocal...microscopy pictures, though, and those can be hard to do as well, you have to sit in the dark forever and take pictures and some people were deeply offended if you listen to music while you do so!  I have no idea what the top panels mean, but I do like pastels (see top panel). For real, really nice work though and something we'll definitely discuss in a lab meeting in the spring semester, for multiple reasons!