Michael Steidel
@michaelsteidel.bsky.social
280 followers 380 following 90 posts
#Cellzome #TeamMassSpec #Proteomics opinions are my own
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michaelsteidel.bsky.social

Weather’s going wild, and now your mass spec data’s a mess too?

Coindidence? Nope!

We reveal how weather-driven air pressure fluctuations impact diaPASEF-based high troughput proteomics - and how to fix it!

Check out our new paper! #diaPASEF #Weatheromics

pubs.acs.org/doi/10.1021/...
Impact of Local Air Pressure on Ion Mobilities and Data Consistency in diaPASEF-Based High Throughput Proteomics
Data-independent acquisition (DIA) on ion mobility mass spectrometers enables deep proteome coverage and high data completeness in large-scale proteomics studies. For advanced acquisition schemes such as parallel accumulation serial fragmentation-based DIA (diaPASEF) stability of ion mobility (1/K0) over time is crucial for consistent data quality. We found that minor changes in environmental air pressure systematically affect the vacuum pressure in the TIMS analyzer, causing ion mobility shifts. By comparing experimental ion mobilities with historical weather data, we attributed observed drifts to fluctuations in the ground air pressure. Moderate air pressure changes of e.g. fifteen mbar induce ion mobility shifts of 0.025 Vs/cm2. These drifts negatively impact peptide quantification across consecutively acquired samples due to drift-dependent abundance changes and increased missing values for ions located at the boundaries of diaPASEF isolation windows, which cannot be corrected by postprocessing. To address this, we applied an in-batch mobility autocalibration feature on a run-wise basis, leading to full elimination of ion mobility drifts.
pubs.acs.org
michaelsteidel.bsky.social
Surprised that u go so low. With EvoSep 24 min method we can load lots more on our Ultra2 until reaching saturation especially with ICC2.0
michaelsteidel.bsky.social
How much are you loading per injection? Is ICC 2.0 enabled on the Ultra2? And which library are you using?
michaelsteidel.bsky.social
True — but the odd part is that the Human Reference Proteome is not really ‘canonical only’. Non-canonical entries from TrEMBL are included, yet the curated SwissProt isoforms are missing default. That’s what undermines the idea of a high-quality reference set.
michaelsteidel.bsky.social
On a separate note: I was surprised to find that none of the non-canonical SwissProt isoforms are included in the official human reference proteome (UP000005640).
Anyone know what’s going on here? 🤔
#proteomics #bioinformatics @pwilmarth.bsky.social il
michaelsteidel.bsky.social
Hey #TeamMassSpec,

Many non-human proteomics studies still search against taxon-filtered FASTAs.

❌ Redundant sequences
❌ Inflated search space
✅ Reference proteomes cut redundancy, improve annotation, and make results comparable.

👉 Time to move beyond taxon filters. #proteomics #massspec #uniprot
Reposted by Michael Steidel
cashwood.proteaglyco.com
Without #2, a lower ion count is needed just to be sure that the full MS range is scanned, but with more accurate ion counts, you can go to the max S/N without losing ions on the edges.
This could also work for the Orbitrap Astral.

Bonus: DIAPASEF on Thermo - patentscope.wipo.int/search/en/de...
Reposted by Michael Steidel
vadim-demichev.bsky.social
With 𝗗𝗜𝗔-𝗡𝗡 𝟮.𝟯.𝟬 Preview (Academia-only for now), we showcase the transformative new capabilities that have been developed in the past months. Download: github.com/vdemichev/Di...
michaelsteidel.bsky.social
Thanks @pwilmarth.bsky.social - also included the less-redundant "one protein per gene" db here ...Has anybody assessed potential benefits of the reduced search space on sensitivity?
michaelsteidel.bsky.social
aaah guess its "hidden" there :)
michaelsteidel.bsky.social
Great thanks! Where can I find the one protein per gene option?
michaelsteidel.bsky.social
Hey #TeamMassSpec,

When you run proteomics on non-human species (mouse, rat, macaque, etc.) — which protein FASTA do you prefer?

Taxonomy-filtered UniProt (all entries)

Reference proteome (SwissProt+TrEMBL)

Ensembl/GENCODE

Something else?
michaelsteidel.bsky.social
Astral Zoom hits >7,000 protein groups & 67,000 precursors — on a 500 SPD EvoSep ENO run.

www.biorxiv.org/content/10.1...
michaelsteidel.bsky.social
Is that narrow windowed thin-PASEF?
michaelsteidel.bsky.social
Sciex in the game.

Pretty impressed by ScanningSWATH data on the new ZenoTOF 8600.

#TeamMassSpec
michaelsteidel.bsky.social
Using 32-Core Threadripper for DIA-NN. Gamechanger for large studies as multithreading is efficiently used. Also in case if several user need to process different jobs at same time.
michaelsteidel.bsky.social
If I got that correctly its not „truly“ open, as only predefined offset masses can be selected.

Have tried Fragpipe22? You can do ion mining from DIA data …
michaelsteidel.bsky.social
..diaPASEF down to 1 ng Hela looks ok (using E.Coli entrapment)
Reposted by Michael Steidel
slavov-n.bsky.social
The videos from the 8th Single-Cell Proteomics Conference (#SCP2025) will be joining this growing YouTube playlist.

www.youtube.com/playlist?lis...