TimsTof Ultra full marketing specs are now live as well!

 I kept looking for this to be up, and it now is. Check out the full marketing dump on the TIMSTOF Ultra!

(Obvious marketing disclaimers, but...I've been told by someone I really truly trust that she/he is obtaining these numbers...)

5500 proteins from 125pg of material... 2k from 15 pg...????

If you haven't been to this awful blog or haven't been able to weather any of my recent tirades in full length, my estimate has been that the amount of peptide I actually get from a cell with 250 picogram of protein is...around 100-150 picograms... 3k proteins is unreal. 5,500 is....

Preprint from Olsen lab -- Astral analyzer details!

Want more details on how the weird new Astral analyzer works? Check out this new preprint! 

https://www.biorxiv.org/content/10.1101/2023.06.02.543374v1.full.pdf

 

SCIEX ASMS Release -- icIEF-MS-coupled to a ZenoTOF!

 

I suspected SCIEX was working on something badass and totally secret. It was all too quiet. I figured it was EchoTOF time or something else...but...even the release has been subtle....unless you're doing antibody characterization (maybe other proteins as well, I don't know).

Our friends at SCIEX have a uniquely weird capillary electrophoresis system or kit that allows them to separate charge variants by their electrophoretic mobility! It's super cool because it is coupled to a UV readout so you can rapidly (a couple of minutes) realize that there is more complexity in your mAB you produced. 

Now....what if you very carefully separated your charge variants....and they eluted right into a 7600 for intact/top down analysis? The drop in complexity in your sample input is going to be so so so helpful. BTW, the 7600 has a huge upper m/z limit. I forget, but it is 40k or 60k or something. Throw in the ability to do EAD/CAD and combinations of the two and you've got a ridiculously poweful solution for antibody characterization. 

I can't find the picture online that I saw, but it is all sort of streamlined together. The box looks a lot more elegant than the name....it's something like icIEF-MS. 

I'm willing to look past the name and think hard about how I'd use this sort of data, and so should you weird biopharma/top down people. 

Waters -- Imaging QQQ!! (Triple Quad) Xevo TQ Absolute!

 

I told everyone not to sleep on the other vendors. This is the biggest ASMS for new hardware advances since I've been doing this stuff.

What are the biggest weaknesses in mass spec imaging? Dynamic range? Limited targeting abilities? 

What if there was an imaging triple quad? 

Check out the Waters Xevo Absolute with DESI! 

What I think we know about the new TIMSTOF Ultra so far!

 

Well... so...if you take the TIMSTOF SCP, and put the TIMSTOF HT digitizer and higher dynamic range TIMS and 

YOU FIX THE AWFUL GOSHDANGED NANOSPRAY SOURCE? 

Did that lady just say 300 HZ????  300 HZ?? I probably misheard her....

For some context, I really have to make my library files for single cell on my TIMSTOF Flex I can't put more than 20 nanograms on the SCP, and 20ng doesn't give you the kind of depth that you get with 400ng on a Flex. But my Flex can't see hardly anything in an unlabeled single cell, so the SCP has to be here.

The Ultra has the sensitivity of the SCP, the dynamic range of a more normal TIMSTOF and it looks way way faster? AND they looked at the worst nanospray source ever designed and tried to do something about it? 

Orbitrap Asstral preprint drop shows how innovative (weird) the new instrument is!

 

We've been had! Clever patent releases and some misconceptions regarding how very sneaky Dr. Makarov can be let us think we were just looking at an Orbitrap with a TOF where the ion trap should be.

It turns out this thing is a whole lot weirder, but I can't spend another 3 minutes on it.

Check out this preprint! 

Label free proteomics R Script Deathmatch!

There's a bunch of R stuff out there and more smart young people who know how to make that awful stuff work. 

How do these packages differ in some of the most important points we care about? 

 

Visualization of diaPASEF methods by Sebastian Paez!

This recent post trying to describe different diaPASEF methods (and missing the one that people are most excited about -- don't worry, I'll come back to it, but I can't lose my job for this blog) got a lot of positive feedback and follow-ups! 

If you missed it, it's here. 

Sebastian took pity on me and my attempts to draw straight lines on IMS windows and made these illustrations. Check out how sick this is.

This is how regular old diaPASEF works when you're running a 6 cycle method! The boxes in the outlines are the 1/k0 and m/z windos that are being isolated in each cycle.

And this is is what is going on when you run the most complicated SlicePASEF-4 (if you download the direct methods from the preprint, I think this is called 4F(?) which always reminds me of all WW2 era Bug Bunny cartoons that I recently watched with my kid in a mix of nostalgia and horror....definitely some concepts in those things that I do not want him thinking are okay or that he should be comfortable with. Yikes. 

Thanks, Sebastian, these totally help me even more to understand what the instrument is doing! 

FlagShip after Flagship in my crystal ball

 

I checked my horoscope this morning and it said that Houston ASMS is going to be probably the biggest year for high end mass spectrometry releases in history. 

I quote from the passage for Capricorn:

Expect at least 3 vendors to release new flagship instruments that will push the boundaries of what you can get from the tiniest amounts of protein imaginable. If you purchased the most expensive LCMS instrument available in December of 2022 at list price, like the wise leaders of a weird little place called Janelia, and still have not hired anyone to run it, you might look even sillier than you do now. 

Then I looked it up and realized that I'm actually a Cancer, but it was too late, I'd already typed this. Who knew that horoscopes were still around? 

Disclaimers are over there --> 

I have no currently valid NDAs and no one tells me anything, this is purely for comedy purposes for a joke about 8 people will get. 

The Orbi-TOF is real! Details leaked on the Orbitrap Asstral!

 

Whether intentional or not (US deadline for S10 high end shared equipment awards is next week, so...) a popular scientific media outlet for chemists has posted a couple of application notes for what appears to be the long-sought Orbi-TOF system. 

Neither application note actually has details on the second mass analyzer (such as whether it is a TOF) but considering that the application note for plasma

Runs a 240,000 resolution MS1 scan and 299 DIA windows in what appears to be PARALLEL????

That's either an ion trap at a resolution of 14 at 200 m/z or 4 ion traps or a respectable TOF. Or....an Orbi-Orbi where your cycle time is 9 seconds or something, but look at the "Loop Time"!! That's 299 scan events in 0.6 seconds? And there is some ridiculously short chromatography in that app note. 

A second app note goes down to 50 picograms cancer cell digest on column and kicks back something like 2,000 proteins.

You might notice there are no links here. I was recently in court as an expert witness and I really truly don't want to be on the other side of things. I'm dying to talk about that case which was an absolute microcosm of everything that is wrong with the current state of mass spectrometry, like why we aren't taken seriously as a distinct scientific field and why so many mass spec labs fail, but I gotta wait out some stuff. 

You'll also notice that they made the great decision of extending the Assend nomenclature. I suspect they'll do something like Hyundai did with the new Genesis brand and we're going to keep going up. Betcha we're just a few years from Assgard. You can find this info yourself by googling the name of the instrument. 

Hopkins 2nd Annual Mass Spec day proteomics overview!

 

(Everyone was sort of on the edges -- and this was Ron Schnaar talking about super complex surface glycans (eeeek)-- the fact anyone is visible is a big thing in my mind! ) 

I started my first real job in clinical chemistry at Johns Hopkins 20 years ago this August (eeeeeeek!) and I've been bumming around the city, hospital and university off and on ever since. For all of that time, there really hasn't been much mass spectrometry here. If you total the hospital system and the university (which are separate entitities) there are just about 50,000 full time employees and 23,000 full time students. That's sort of big for a school that has never really had even a Metabolomics core. 

SO...the fact that there were two rooms full of posters with a lot of people I DIDN'T EVEN KNOW showing all sorts of mass spectrometry data is super exciting. 

Highlights for this reader base? 

Chan-Hyun Na showed some great stuff on how his group is finding non-canonical peptides in Alzheimer's models. That is, of course, a protein disease that genomics isn't all that helpful with. While I'm a huge fan of the Steen lab work on this topic, I had no idea they were doing this super effectively across the street. 

Stephen Fried doesn't just do really mass spectrometry to solve fundamental questions about the earliest stages of enzyme evolution (cool recent paper here). He demonstrated some applications of similar approaches to understand neurological decline. 

Rahul Bharadwaj showed some work with Bob Cole's core (I think Lauren DeVine will have a poster at ASMS showing this data, it was too popular for me to get to it), super ridiculously small laser capture microdissection samples + high plex TMT proteomics (MS3 based quan, I think, I can't always read my notes) from people who were so amazing that they left their brains to the Lieber brain banks. Super cool stuff. 

Ron Schnaar studies the most frustrating looking cell surface glycans of profound medical importance through photoreactive probes his team synthesizes themselves. He made a really solid argument for why intricate biological pull-down studies benefit from the super high accuracy TMT multiplexing you get on the Orbitraps largely designed for that purpose. When your biology and the molecular aspects of your experiment are super complex, it is nice to have the best quan on the back end that you can get (and the internal QC aspects of multiplexing). 

Josh Smith talked about his continued work with adductomics -- there it is -- I was sure I had a blog post on a paper of his from a few years ago. If you aren't familiar, it is cool and scary stuff. Actually, that paper was on targeting -- here is a very recent JASMS study doing it untargeted! 

That was only half the day! There were great posters (Bob O'Meally is making custom SureQuant panels, check out his poster if you're in Houston, it's really cool) and the rest of the day was imaging and them small molecules and there was even a single cell ICP-MS (accurately measuring metal concentrations in one cell at a time). Super cool day. 

Top down proteomic analysis from a single muscle cell!

 

Woooo!!  While most people are trying to get usable shotgun data out of their first actual single cell right now, there are a couple groups out there trying to do the impossible -- TOP DOWN proteomics from single cells. 

During the Proteomics Show Road to Chicago we talked to both Alexander Ivanov and Ying Ge and I was surprised to find out they were both doing it -- and both have published now! 

You can check out the new study from Ge lab here! There is an earlier preprinted version if you can't figure out how to log onto PNAS through your library....

How the f' do you get proteoform analysis from a single cell? The technology they break out here is worth the time to read through. They go to capillary flow rates (2uL a minute) and use the funny 8 nozzle source from MnESI that Simion Kreimer is using out a Cedars on their TIMSTOF SCPs. Then they use the MAXXIS! That's the ridiculously tall TOF that gets 80,000 resolution across the entire mass range. I've only seen two in my life. One was in the Bruker demo lab and the other one (which, quite honestly, might be the only one that has ever been sold) is in Madison.

Turns out you can get over the sensitivity hurdle for the device by using 8 different 10 micron ESI emitters. Also, the muscle cells appear to be sort of huge, so that probably helps.

However, they resolve proteoform level differences -- between different muscle fiber cells (!!!) in proteins in excess of 200kDa (!!!) I've only resolved a couple of proteins in my career above 200kDa and typically at the sort of resolution to say "yup, it's probably about 10kDa bigger than you said". What they've done here -- alone -- sets this study up in rare air. To do it in a physiologically relevant context? In single f'ing cells? 

Huge props to this team. 100% recommended read that sort of resets some of the boundaries of what we can do today if we're willing to think outside the box. 

m.2 SSDs can double HTRMS data conversion rates vs eSATA SSDs!

 

(Image above stolen from this article intended for people who edit video) 

I recently had a tirade about making these new fangled DIA data processing software runs faster. It was focused on SpectroNaut, but I think it will carry over to other tools equally. 

Honestly, it was a desperation thing for me because we were bottlenecked like crazy at the data processing things and -- things don't seem nearly as desperate now.

These numbers this morning are the most striking to me. 

The first thing SpectroNaut does is convert your whatever it is data type to the HTRMS data that it likes. 

Check out how long it takes to convert 55,000 diaPASEF MS2 spectra from one file on a Samsung Pro SATA SSD drive

VS comparable sized files on the Samsung Pro M2 (plugged directly into the motherboard) drive!

The HTRMS converter doesn't keep the processing time information forever in the GUI, so I had to just pull the last file I ran off of the SATA SSD and the next files which I ran exclusively on the M.2 drive.

When you've got multiple grant proposals due first week of June and your front office wants them turned in Wednesday, saving 3 minutes each on 90-ish files on just the very first step of your processing pipeline can seem like a dogsend. 

FAIMS improves (?) the identification of host cell proteins in antibody drugs!

 

Antibody based drugs are developing at an absolutely phenomenal rate. This new study throws some caution cold water on the party by digging deep into the fact these antibodies are coming from cells and that sometimes other proteins come along for the ride. 

It isn't my job to weigh in on whether this is actually a big deal or not, but I hope that it isn't. Or...I hope that if it is, it's easy to fix. 

For some context from the mass spec side, if you have to get out an Orbitrap Eclipse and run a 2 hour gradient at nanoflow rates (IonOpticks 25cm!) using the ion trap so you have the highest sensitivity on your MS2s and then put on a FAIMS source to get a couple dozen peptides I'll go out on a limb and say -- there ain't much there. You have to be talking about a tiny tiny fraction of contamination, right? 

Interesting to think about, and I'm cautiously optimistic that this is actually saying very positive things about the mAB production industry. However, I guess if that mAB came from a hamster ovary cell and I was deathly allergic to hamsters rather than just grossed out by them, I'd probably want to know those 60 peptides were there? 

Put a question mark in the title in case this is something like part per Trillion detection and it isn't remotely biologically relevant and is just annoying to people making antibody drugs. 

PipeR - Proteomics to actionable, accessible and expertly designed output for physicians!!!

 

WOOOOOOOO!!! I'm leaving this here, then checking my calendar and seeing if I can install this without help. And then getting help. Or....doing whatever my calendar tells me to, then doing the things above. 

Wait. I don't even have to install it to try it out. 

Check out the PipeR shiny.io interface here!

Click to enlarge, or go try it out yourself lazy bones. Pick the patient (1), pick the clinical phenotype you're interested in (3), choose how you want the data plotted. 

Wait. What was that panel again? BOOM! Click the panel details tab! It's like these people have called a sleepy resident at 3 am to deliver a crazy blood potassium level and this poor person who has been on the floor for 28 straight hours -- is like "remind me what the normal range is again, please...?" If you're the clinical chemist who has never had to pull more than a 16 hr shift on a holiday it's easy to be sort of dismissive of the physician, but there are reasons most people (even in the US) aren't allowed to work more than 16 hour shifts. You don't get smarter at hour 17. Reminders are embedded in reports that go out for very good reasons. 

I can't over express how much I love this whole idea!