Wednesday, October 5, 2022

FAIMS settings for ultralow (and ultra-ultra low flow) proteomics!

 


Do you have one of those big FAIMS things killing dewars of N2 in your lab...or...

...well...

...in either case, if your question is something like -- "Hey! What buttons would I press if I wanted to analyze <1 nanogram of peptide (why would you want to do that...?)?" And or "what FAIMS settings make sense for ultra low flow analysis?"

Check out this study where they worked it all out!



Tuesday, October 4, 2022

Study of DNA polyploidy cells is really a nice analysis of RNA-Proteomic correlations!

 


I started reading this because cancer polyploidy is really strange and very cool (honestly, I thought this was going to be background so I could better understand a recent paper out of Josh Elias's lab). Turns out the two things are only loosely related, and if I sort out the latter enough that my notes make sense, I'll come back to it later. 



If you aren't familiar with polyploidy, DNA divides in these cells, but the cells themselves don't. And some evidence supports them as being the origin of metastasic lesions. BIG DEAL. 

While I'm trying to force some new concepts into my tired old brain, there is a lot of this paper that is less difficult to grasp. 

Including some really exhaustive analysis of RNA to protein abundance correlations! No surprise, they aren't very good (the "correlations") but there is tons of great new stuff in this study, so the orange publish button will probably get hit. 


Monday, October 3, 2022

qPTM -- A nicely organized database of PTMs in some model organisms!

 


This wasn't the resource I keep dreaming of finding, but I do still like it


I'm not sure how this is better than PhosphoSitePlus and whatever the other thing is called that you can link to from UniProt now, except in terms of how it is organized. 

Think Mondays are a stupid day to read and just want to type something into a box? You can access this resource here. 

http://qptm.omicsbio.info/

Sunday, October 2, 2022

Saturday, October 1, 2022

Compare MS2 2.0 version 2 (dos, zwei, ni, tva, 'er, meiji, do, two, II)

 


Straight up, this kind of seems like some sort of crazy street magic stuff, but the nitpicky nerds that get to review for JPR let this one through, so I'm going to leave this here so I can look at it later. 


Weighting mass spectra -- across species? What the eff? 

Friday, September 23, 2022

UCDavis 2020 Proteomics Shortcourse is online?!?!

 

I'm always looking for great tutorials for the "Resources for Newbies" page over there --> and I just found out that the legendary UCDAVIS proteomics short course from 2020 is 100% online! 

I'll obviously add this to my list of resources but you can check it all out here

Thursday, September 22, 2022

Time to toss that nanoLC off the roof of the parking garage!

 

I've tried to hide my hatred for nanoflow liquid chromatography for over a decade now, but rumor is I haven't done it very well.

For more motivation to carry that thing up to the roof for real, I present a really nice study in JPR. 


In this work they pit a nice shiny Exploris 240 system with capillary flow (1 microliter/minute, which I'd still consider nanoflow vs microflow (50uL/min) in a variety of different contexts across multiple sample loads and total experiment acquisition times. 

It is largely what you're expecting --- peak shapes are better at higher flow rates, and lower flow rates do better with lower sample loads. 

Where all the work this team did really shines is in where you can get close to matching the coverage at these high flow rates vs low ones and you find these points where it seems pretty darned smart to use the 50uL/min. 

For example, for 15 minute LC runs, 2ug or 5ug sample load is pretty darned close at 50uL/min or 1uL/min. You could argue that was probably just too much sample for the little column used for the 1uL/min gradient, but still...we're talking about almost 2,000 protein IDs in 15 minutes! That's legit. 

Wednesday, September 21, 2022

Advances in Proteomics and Metabolomics 2022 by Tech networks!

 


If COVID-19 did anything for the world besides getting rid of a surprising number of people with the last name "Orsburn" it did, at least, bring digital meetings forward. So many opportunities to watch cool talks! 

I was just tipped off to this one and it looks great

Tuesday, September 20, 2022

Ubuntu summer school -- 5 days of proteomics in Antarctica!

 


Someone sent me this link to this very slow loading website where a ton of great speakers are teaching a summer proteomic school where there are penguins! As my public education didn't include a geography class and under the registration page there is a picture of an amoeba?....?...


....I'm going to make the uneducated guess that this meeting is in Antarctica. That's where my grad school roommate worked for 7 years but now got downgraded to working in the middle west somewhere. 

For real, if you want to see penguins and spend 5 days with some of the top proteomics researchers in thw world, registration is only open until October 14th! 

Monday, September 19, 2022

Study protein turnover IN VIVO in PEOPLE with DeuteRater-H!

 

"...because serial biopsies in humans are impractical..." (yeah...maybe a stronger word belongs at the end, but the point stands) it is really hard to truly understand in vivo protein turn over in humans.

So....what if you could talk an IRB (and some undergrads?) into having the latter drink deuterated water every day for a while and provide you with saliva serum and muscle samples here and there? 


Well...you'd have this study

Samples were analyzed on a tribrid Orbitrap system and to work out this new metabolic model, you'd need new software and you can get that here

Wow, right?!?!

Sunday, September 18, 2022

3 upcoming US HUPO Webinars!

 


Until recently I kept forgetting that US HUPO was a thing, it was even just outside of Baltimore a couple of years ago and I just forgot it was a thing until cool people started texting me to see where I was. This isn't a dig on US HUPO, we just got into some totally f'ed up traffic last week because I forgot that American football is still a thing and right after a match we were on the wrong side of a stadium from home and it took a really long time around people who appeared to be driving somewhat or incredibly impaired. 

Well, US HUPO IS a thing and they are putting on all sorts of great remote learning things all the time! There are 3 great ones of note in just the next 3 weeks! You can check them out and register here

Wednesday, September 14, 2022

16 proteomics sample prep methods head-to-head! The quest for the universal method!

 

If LCMS proteomics is going to scale up to the levels of new proteomics technologies that are willing to compromise specificity for throughput, we need to trim down the 4,512 different sample prep methods each lab has sitting around in stacks. You can't tell make a sensible argument that about 4,400 of them are unnecessary. 

What we need is some people to do the most boring studies imaginable just to get them out of the way. 

OMG. From the pit of my heart, I can't thank these authors enough for this study. It should be referenced by everyone in proteomics all the time. "What do you think about method A vs method B"? I have a hunch but nothing robustly tested but --- 


-- these awesome people tested them all! (Well...not all 4,512....but a whole ton of them!) 

Sure -- number of proteins are one metric -- but they ALSO break down the relative costs of the different methods! Proteomics sample prep has historically been pretty cheap from a reagent perspective compared to DNA/RNA assays that still require hundreds of dollars in reagents. But when you're setting up a 100 sample or a 4,000 sample study, those filters or traps and C-18 spin columns (and TRYPSIN) costs don't seem that irrelevant anymore. On a big study I recently realized that the 1:10 trypsin ratios recommended for some commercial kits was going to blow through every vial in our -20C in very short order. This group considered these factors! 

What wins? Nah, you gotta read this so you see how awesome it is. And you've got to remember to cite it later! 

Tuesday, September 13, 2022

No more Orbitrap Assend Leaks! Here is the full marketing dump!

 



As I'm looking over mostly completed blog posts and wondering why I didn't submit them, or removing completely unrelated gifs that make less sense than the ones I typically insert, I refreshed on the Orbitrap Assend official site and found tons of cool new information! (Working at a level that probably doesn't make sense in any possible way right now). 

This is old news for any of you lucky people who were in Maastricht, but for those of us waiting on the edge of our seats for every little Assend leak to trickle out, this big drop of information finally puts the hardware in perspective. 

There is a reeally cool little video at the bottom of the page with some gripping piano music that shows the whole ion movement from beginning to end for a typical data dependent experiment. 

(Please see disclaimers over there --> as always, I'm just trying to help get information out there!) 


Based on the architecture itself and what has been publicly released, the performance gains reported by Gygi Lab and Coon Lab are much better than I was expecting. This suggests to me that the obvious new fetures added (like a whole new multipole) aren't the whole story. There is likely some additional efficiency gains here and there. 

Honestly, on paper, I never expected the Fusion 3 Eclipse to be a big step up from the Fusion 2 for data dependent shotgun proteomics (I thought it was just for intacts and they'd kind of forgot about the DDA crowd), but when you actually process the data I have been impressed by the step up. It is easy to forget how complex modern instruments are and how little much little improvements like slightly reducing a gating time can lead to big improvements. 

Monday, September 12, 2022

"Next gen" proteomics deathmatch is published! Who cares about specificity?

 


Wow! Look at that, O-link looks great, see how it matches an error-prone protein quantification assay that we typically use when measuring as inexpensively as possible is the most important and accuracy is secondary? 

Oh. I particularly like this one as well! 


For my entry in the "most times I have literally laughed out loud while reading a peer reviewed publication while operating on less than 50 hours of sleep in an 11 day grant writing stretch "  I humbly submit --


Would you like to see my favorite funny part of this? To be fair, fate transpired to have my first solo grant application ever overlapping with 3 sets of really great journal reviewer comments all being due at the same time and my watch kept giving me this angry frown for my sleep tracker -- and I have had some sincere concerns about the quality of my driving on the beltway this week, so this might not be as funny next week as it is right now. (There's the period key! Looks like I forgot where it was for a second!) (Please, no real concerns about whether I'll die on the strech of my commute that we in the world's greatest city refer to lovingly as "THE THUNDERDOME" because, on a 5 year average approximately one person will enter per day who will not leave, when I'm really sleepy I drive my old stick shift car and it drives itself.) 

Check this out and tell me it isn't funny. 


Highlight 1 (my interpretation): Man, we're way more streamlined than LCMS! You should use our "next gen" technology if  you don't mind that what we're quantifying isn't SPECIFIC to the protein that we think it is. 

Highlight 2. Independent verification of our performance relative to a gold standard (I assume LCMS) for each of the thousands of proteins on these platforms is expensive and it will take a long time!

What comes to mind when you think about SOMASCAN and O-Link investors who are jumping on the proteomics bandwagon and are really excited about how much money these companies are spending on advertising their products because proving that they work is going to take a long time and will be expensive? 

Personally, I imagine that they're very tech savvy people who bring broad expertise in from whatever field they come from that readily translates to a deep understanding of any new technology!  

Time to change the topic to something completely different.

I don't think I've ever enjoyed a movie with Jim Carrey(sp?) in it enought to finish it. Even when he was seemingly in every 4th film in theaters I just found him completely unwatchable. However, I found this gif somewhere and I don't even know what movie it is from, but I have worn something remarkably similar to what he is wearing in this gif below. 

Wait. Is this the right blog? I should get some sleep. 

Saturday, September 10, 2022

IMSC2022 Final guest blog recap by Dr. Nick Riley!

 

Pop to part 1 or part 2 here.

IMSC 2022 Part 3: Wednesday to Friday

Wednesday, August 31, Day 3

Science Summary: It is difficult to pick between the best plenary/award talks of the week, but the Wednesday morning talks from Curt Brunnee Award winners Livia Eberlin and Erin Baker and inaugural Jochen Franzen Award winner Shane Ellis might have taken the cake. Following those, I saw great talks from Tami Geiger and Hem Gurung in the Translational MS/Cancer and Immunology morning session, and then chased talks between the Top Down and Imaging MS sessions after that. Wednesday saw a change over of posters (M/T vs W/Th groups), which brought more great poster discussions after a Thermo lunch seminar from Daniel Lopez Ferrar and Kay Oppermann about the AccelerOme. I spent most of my afternoon time in the second Glycomics and Glycoproteomics session of the week that featured interesting work focusing on structural characterization of both glycans and glycoproteins from Javier Torano and Cathy Costello, among others. I also was able to catch some cool real-time ID/quant work from both Gad Armony (Wessels group) and Chris McGann (Schweppe group). One of the highlights for me was Noortje de Haan’s talk in the Glyco session that showcased how glycogenomics and MS can work in tandem for O-glyco studies. If that sounds interesting to you, check out recent work from the Copenhagen Center for Glycomics.

 

Social Summary: Navigating the bus system in Maastricht is relatively straightforward, unless you assume there is only one train station. I confidently claimed a bus was going to the train station and led a group to the lesser-known, non-central train station several miles north of the city by mistake. I guess this is the price I pay for trying to only rely on free wifi at the conference center and hotel instead of double checking these things in real time with cellular data (which I eventually turned on). My bad, folks.

Culinary/Cultural Highlights: Erin Baker gave a nice (terrifying?) summary of how PFAS is everywhere, including our plastic food wrappers. It made the loud crinkling of plastic wrappers of the pre-packaged food during the lunch seminars sound that much more noticeable with a few more side glances than previous days. On a different note, the non-plastic-wrapped snacks and refreshments during the breaks and poster sessions seemed to only get better with each passing day.

Thursday, September 1, Day 4

Science Summary: A consistent theme here is the excellence of the morning award talks, and the presentation from Thomson Medal winner Vicki Wysocki was no exception. I unfortunately had to step out and miss the Thomson Medal talk from Lidiya Gall, but I was glad I at least was able to hear her present during the FeMS workshop on Monday night. Morning talks in the Biosimilars, Biobetters, and Glycoengineering session were mostly antibody-focused, with my favorite being the multi-protease approach to de novo antibody sequencing from Weiwei Peng (Snijder group). I also liked Johannes Helm’s (Altmann group) talk about PGC for isomer-specific N-glycomics and Mike Gross’s talk about different footprinting techniques, especially NanoPOMP, for membrane proteins. I spent my afternoon session in the second Young Mass Spectrometrist session, and each talk was impressive. If you are looking to know where the next generation of MS research is heading and you haven’t seen work from Karina Gonzalez-Estanol, Na Wu, Lieke Lamont, Guinevere Lageveen-Kammeijer, or Andrej Grgic, I highly recommend you check out their work.

Social Summary: The conference closing event was a fitting celebration of the science and scientists at the meeting. Set along a small harbor in Maastricht, it was an outdoor setting that underscored the good weather and fun city that IMSC 2022 enjoyed. I think this tweet summarized things nicely:


(Note, the time stamp is for my current time zone, not the local time zone where it was tweeted – I think.)

Culinary/Cultural Highlights: The couponed food was all pretty enjoyable, with one notable exception. Perhaps the Europeans will take issue with this assessment, but the trout paste “delicacy” was an acquired taste that several us had yet to master. Luckily, the other good food and non-couponed drinks helped clear the palate.

 Friday, September 2, Day 5

Science Summary: Similar to ASMS, if you stick around until the last day at IMSC, you are likely to enjoy some excellent science without as crowded of rooms. Still, I thought Friday was relatively well attended considering the jam-packed week it had been. Morning sessions included really interesting talks in the second High Resolution MS session that included everything from lipid nanoparticles to 18-plex TMT work on the Orbitrap Ascend, and the PTM crosstalk session and the second Imaging MS session also had full lineups of great presentations. The afternoon included a closing plenary and outlook for the next IMSC in Melbourne. To summarize the entire IMSC 2022 experience, it was quality science and quality social interactions from top to bottom. As I said in the first post, IMSC is now a conference that will consistently be on my radar.

 

Social Summary: I had a list of people I had wanted to catch up with or meet throughout the week, and a thinned out crowed on Friday morning helped facilitate a few meetings I was glad to have. I never really thought about it this way before, but for younger researchers who may want to connect with more established folks, these last conference days when everyone is a little less flooded with interactions from all sides might be useful. That said, beware that fatigue is real, and not everyone will be ready to have in-depth discussions.

 

Culinary/Cultural Highlights: The snack during the breaks between presentationso n Friday was vlaai, a Dutch pie-esque pastry, and it was amazing. Keep it on your list of food to try if you make it to the Netherlands.


Friday, September 9, 2022

IMSC2022 -- Part 2 by guest blogger Dr. Nick Riley!

 For the first part of Dr. Riley's recap of IMSC2022! 

 For the final part, click here!

IMSC 2022 Part 2: Sunday to Tuesday

For frame of reference, days started at 8:30a, lunch was generally 1-2p, even workshops started at 6p, and most nights did not end until well after midnight for those who wandered into the city center. So yes, the typical conference schedule.

Sunday, August 28, Day 0

Social Summary: The opening ceremony kicked off with a short introduction of talking that was punctuated by a 15 min samba drum performance. Yes, really. The group called Segura had interspersed throughout the crowd and slowly made their way to the stage until the full ensemble danced to their drum beats. It was very cool and unexpected, and it helped some bleary-eyed travelers fighting jetlag (read: me) wake up and enjoy the opening lectures.

Culinary/Cultural Highlights: The opening reception was a great first connection point to talk with folks. It went a little late to be able to grab dinner afterward (things close early on Sunday nights in the Netherlands), but lucky a local döner stand was there to make sure several of us got food. If you haven’t enjoyed a döner kebab before, imagine the Turkish version of a gyro in a pita pocket instead of a pita wrap. They are always a highlight of traveling in Europe.

Monday, August 29, Day 1

Social Summary: The FeMS Workshop led by Anne Bendt (a found of FeMS) and Purva Kulkarni included talks from Jenny van Eyk, Manfred Wuhrer, Berta Cillero Pastor, and Vicki Wysocki (who pulled double duty and came to present after leading the concurrent Native MS Workshop). Erin Baker and Cathy Costello sat right behind me in the workshop and offered some highly useful insights, too. The drink hour after the workshops helped everyone connect and find dinner plans after the first full day of the conference. Instrument companies did not have hospitality suites per se, but instead claimed local bars in the city center as their hang out. I did not hear any complaints.

Culinary/Cultural Highlights: Public service announcement: burritos in the Netherlands != burritos in the Bay Area (or the US in general for that matter). Still, they were open late after the workshops, so they were a welcomed sight. They come with an optional side of fries… not the typical burrito accompaniment, but they were delicious nonetheless and stole the show from the burrito.

Tuesday, August 30, Day 2

Social Summary: More evening workshops kept some attendees engaged, but I, for one, could feel the slightest air of fatigue setting in after the afternoon talks wrapped up. Regardless, it seems as if everyone collectively found their second wind after dinner because the city center played host to plenty of IMSC badger-wearers late into the night.

Culinary/Cultural Highlights: A group of us randomly stumbled into a wonderful Greek restaurant that was an unexpected treat. Also, the local Stadtbrouerij made for an enjoyable aperitif. On another note, Maastricht must host a decent number of conferences or events, because the bus drivers were not phased in the slightest by 50+ people piling onto the bus at the conference center stop, all of whom were riding for free as long as they had their conference badget (thanks IMSC organizers!).

 

Thursday, September 8, 2022

Nice new review of machine learning in proteomics!

 


We're at a point where if you aren't using machine learning or deep machining or intelligent artificialness in proteomics data processing, you're on the outside looking in.

For a great recent review of the topic check this out


Wednesday, September 7, 2022

IMSC2022 Recap by guest blogger Dr. Nick Riley Part 1

I found the password to the blog! 

For the first post, I'd like to talk about IMSC2022. If you had COVID and it affected your memory, 

IMSC stands for the Independent Mystery Shopper's Coalition, and a lot of people went to Maastricht recently to see the release of the Orbitrap Assend there!

If you want to learn about it, don't go to the homepage, you'll want to put the year of the conference before the .com (https://www.imsc2022.com/). Don't leave out the year, for real. 

One person who did go to Maastricht was Dr. Nick Riley. Nick is a postdoc in the Bertozzi group at Stanford and he volunteered to provide an on-the-ground perspective of this conference. 


Somehow, Dr. Riley found time to write this between his trip from wherever Stanford is to Maastricht, where he was a keynote, before he raced back to a beautiful wedding! Not one he was attending, HIS wedding. So I doubled down, and found where the new password to this blog was so I could post this. I wish the photogenic couple all the best. 

Enough words from me: Take it from here, Dr. Riley! 

IMSC 2022 Part 1: Overall Summary

I had the privilege of attending the International Mass Spectrometry Conference this past week in Maastricht, the Netherlands. The vibrancy of this year’s IMSC is hard to capture in words, but I will try to do as much here. It was a thoroughly enjoyable experience and is probably in the top 3 for conferences I have attended thus far in my decade in research. A mixture of great science, wonderful personal connections, and a highly enjoyable European backdrop made for an unforgettable meeting. Thank you to the organizers (Albert Heck, Ron Heeren, Manfred Wuhrer, the Dutch Society for Mass Spec [NVMS], and all those who helped otherwise) for all of your efforts make IMSC a success. Here is my best attempt to distill the meeting down into a few “short” descriptions for those who could not attend.

Some first thoughts

The size of IMSC struck a balance between Gordon or Keystone Conferences and ASMS. There were approximately 1300 attendees if I remember correctly, with a majority (unsurprisingly) from Europe. It was nice as a US-based attendee to hear from and interact with many brilliant scientists who I only previously knew from publications. This different cross-section of science I was exposed to the past few days was a healthy reminder that ASMS, as great as it is, still only captures/emphasizes a limited scope of our field.

The general daily structure of the meeting followed a common pattern. Each day started with morning award presentations that were the only event on the schedule. This was a good thing, because they were all outstanding and deserved full attention. My notes from each of them include as many thoughts about life/research advice as they do about the science that earned the awards. Those talks were followed with a very welcomed coffee break that often featured baked goods and facilitated a lot of science cross-pollination. Concurrent oral presentation sessions followed the coffee break, then a lunch hour where vendors had presentations to attend, a poster session to start the early afternoon, later afternoon concurrent oral presentation sessions, and then evening workshops. All poster sessions and breaks featured access to gratuitous amounts of coffee and tea, which really helped this jetlagged American survive. Overall, the quality of both oral and poster presentations was stellar. If there was one topic that had the most representation, it was probably mass spec imaging, which should not be surprising given the proximity of the M4i Institute, but there was a good mix ranging broadly from instrumentation and applications. I’ll touch more on high-level details from each day in two subsequent entries.

 Instrument companies and other industry partners were present and accessible, but not dominant. Perhaps it is my own bias that I feel this way, but the user meetings and hospitality suites at ASMS make the instrument companies feel more omnipresent at ASMS than they did at IMSC. That is not a criticism, mind you. I like the excitement that comes with the presence of instrument companies and industry partners at ASMS. That said, it was a refreshing change of pace to have it be less of a focus. The most exciting instrument development of the meeting (to me) was Thermo’s release of their newest Tribrid platform, the Orbitrap Ascend (but I fully admit that I have a Thermo bias in my instrument usage history). The main presentation about the Ascend came during a lunch meeting at IMSC, which was pleasantly lowkey relative to the fanfare that can accompany instrument releases at ASMS. Perhaps the oddest aspect of the meeting was the Innovation Lab that happened in mid-afternoons, where industry partners took to a catwalk in the middle of a poster hall to strut back and forth while presenting for 15 min short talks (I’m not exaggerating). The Innovation Lab was “intriguing” if nothing else and did succeed in engaging a decent number of attendees, but it would probably be the event I would recommend leaving out in the future.

 Maastricht seemed to be an ideal setting. As I spent my first hours walking around the city before the conference started, all I could think to myself was “simply charming”… which was weird because that is not a phrase that exists in my world very often. A small-ish university town, Maastricht was energetic and easy to navigate. It probably did not hurt that the students had returned and were starting classes soon, lending a pervasive energy that spilled over to us mass spectrometrists. One of my key takeaways was how valuable walkable cities and good public transportation infrastructure can be for fostering connections between scientists at a conference (and beyond, of course).

 If I was forced to offer constructive criticism, the only thing I found myself wanting was recordings of the presentations during concurrent oral presentation sessions. Many of us scurried between conference rooms to catch back-to-back talks, and I missed as many good talks as I saw merely due to scheduling. (Luckily the conference center had a relatively compact footprint.) This is always the struggle with conferences that have too much good content, and it would have been nice to know I could go watch what I missed. To give a quick snapshot of what I mean: I had the honor of presenting in a Young Mass Spectrometrist session and would not trade what I learned during the talks during that session. However, I missed what I know to be fantastic talks in the Protein-Protein Interaction session, Structural Biology session, and “Beyond Mass spectrometry: Making MS Obsolete” session. Such is the burden for attending a conference with high quality science at every turn. On the flip side, the lack of recording meant I was as tuned-in as ever for the talks I did see.

 The last thing I’ll say for now is that if ASMS is a marathon, IMSC was an Ironman. The extra day of events relative to ASMS (IMSC was Sunday through Friday) coupled with a 9-hour time difference (for west coast folks) and unfettered access to European beers made for an exhausting, albeit exhilarating, week. I would not have had it any other way. IMSC was announced to be in Melbourne, Australia in August 2024 (cc: Gavin Reid) and Lyon, France in 2026 (I believe). My strong recommendation: do yourself a favor and circle both dates on your calendar in bright red like I have.

Tuesday, August 23, 2022

Fusion 4 Details leaked! It IS real! Introducing the Orbitrap Assend!

 

For legal purposes, we'll be clear up front that this is up on this website. No insider information here ever. Who would tell me things? 


What do we know so far? A little. 

The schematics appear to show two separate ion routing multipoles, which can only increase parallelization capabilities. Oh cool! A full document with schematics is up on the website link above now! 


Some pretty amazing numbers here. The optional HighMassRange option goes to 16,000 m/z. That might be the highest number outside of the E+EMR or UHMR (I forget that latter's upper number). Orbitrap is talking about 45 Hz @ 7,500 resolution? 

The autoready ion source appears to be a self-calibrating option, which would explain some of the marketing stuff we're seeing cool videos of. The document says you can auto schedule calibrations in your queue which is an Orbitrap first.

For anyone hoping that we'd see a TOF back there, you might be disappointed, but it looks like the ion trap was again tinkered with to improve UVPD, and PTCR. 

It seems like full details will release at IMSC which just wasn't on my radar this year and I don't know where I had that cognitive gap to not even consider putting it on the calendar. For the first big launch from this vendor in quite a long time, and a really impressively professional series of marketing hints and links, I suspect it will be quite the official launch. Sad I'll miss it, but hopefully some data will leak and more will show up at that little conference in rural Mexico in December. 

Monday, August 22, 2022

Super expert TMT Post #2 : How to mix plexes by Alejandro Brenes!

 See post #1 yesterday, but if coming from somewhere outside this blog. Here is the link


This original paper has appeared on the blog at some point, for sure, maybe twice because it is amazing. But it should be posted here again because it is such an important topic


The original paper provides advice for how to mix different plexes, what to definitely avoid and has all the math to back up the fact they are not making this up! 

Why post it again? When this paper was submitted the TMTPro18 reagents weren't yet out. Rather than making you wait for him to wrap up his thesis and find it (I'm sure it will be worth reading!) he made the updated diagrams available for these reagents! 


Higher resolution is available here (I can't embed twitter's new .jiffyjaff format or whatever it is in blogger. Off topic, but Amazon now uses a new image format for everything, including your personal images if you use their cloud backup thingy. I can't embed those either.)

I bugged him about my own personal issues with medium resolution instruments and multiplexing (on the TIMSTOF and 7600 we can only 10-plex using 126, 135 and 127-134 and he made this as well (you can find the higher res in the same Tweeter thread in the link above).  


It is funny because just a couple of years ago when we were limited to 10-plex on ultra high res instruments, a lot of people independently decided that combining plexes wasn't worth it. CPTAC plowed on combining plexes leaving the quiet implication on the table that maybe we should read their papers in their entirety rather than just working through to figure out just how much one of these amazingly expertly analyzed tumor samples cost on average. (Whoever said this last round cost $15,600 per proteome was probably mediocre at arithmetic). 

Whether it was informatics that needed to advance or study design, combining plexes is now common. There are absolutely clearly pitfalls when you do this, but if you've never set one up before this is THE paper you should read first. 


Sunday, August 21, 2022

Super expert advice for TMT post #1: TMT Channel Crosstalk by Phil Wilmarth

 

The next two posts will blatantly steal other people's work who are just trying to make sure that we're mixing TMTPlexes and considering all the proper variables

For post #1: This link will direct you to a real proteomics science blog everyone should read!  

Saturday, August 20, 2022

Full structure T-Cell receptor ligated by MHC!

 

It's easy to forget about protein 3D structure sometimes when you spend your week trying to tell linear peptide sequences apart. In the end, however, a lot of actual protein chemistry is "if this thing is open then this other thing fits in it", right? 

This stunning new study demonstrates one of the most critical parts of how MHC binding to T-cell receptors works and angles are super important! (CryoEM is getting better all the time!) 


What comes out of this study that is really interesting is how critical the base of the protein complex is and the role that lipids clearly play in facilitating the interaction between the complexes. 


Friday, August 19, 2022

Targeting protein splice variants -- the definitive guide!


This might not be the most exciting thing to absolutely everyone, but I really like this handy step-by-step protocol for targeting those splice variants that we like to ignore in global proteomics data. 
 

Loads of proteins work by cutting other proteins and splicing them together into new forms with altered or completely new functions. If that's what your collaborators are actually interested in, global proteomics can be a pain in the butt. I've had a PC just about bricked for 5 days processing 170 TIMSTOF files with DIA-NN and found out during the data review that my spectral library didn't contain the single peptide that my friends actually cared about. Considering that relative costs per sample of the $1.3M list price TIMSTOF Flex + MALDI 2 that I used for 2 weeks of data acquisition vs. our very nice, and definitely not $1.3M list price Agilent Ultivo (the world's largest HPLC isn't nearly as large in person as it appears in the ads). You might argue successfully that had everyone been using the exact same language, I could have very easily targeted the single peptide variant...

(unrelated, but funny)

...to be fair, however, finding the sequence variant was NOT FUN. It was not listed in UniProt or NCBI as a peptide isoform variant, so I did have to find the nucleotides and work out the codons and make my target peptide, and send it to PROSIT to make my targeted list (I am specifically just reprocessing the diaPASEF data for that variant now). Skyline LOVES 170 diaPASEF files, btw. I expect to have quan well before US HUPO. 

And this is why I liked this new protocol. There are multiple tools listed in the review for converting DNA sequences to splice variants and super easy instructions (and pitfalls) for targeting these in Skyline. Given that these authors say that OVER 90%!?!?!?!? of human genes go through some sort of splicing!?!??!?! I think this protocol might go into the permanent folder on my desktop that just says SUPER USEFU (maybe if it was lower case the L would fit, but I think I need the caps here).

Thursday, August 18, 2022

Make optimized windows for diaPASEF (& for phosphoproteomics) with pyDIAID!


As I might have whined about in the past, making pasefDIA windows can be some tricky business. We gave up and had an expert come in, load up his method, and we just run that one. Some other group wasn't satisfied with that method and made a program to make optimal methods!  


The program is really straight-forward to install and run on what appears to be every operating system and you can get it here.

The dividends for optimizing windows for phosphoproteomics shown in the study are seriously impressive compared to just running a canned method.