Michael Steidel

Which LC & Flow?

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

How much are you loading per injection? Is ICC 2.0 enabled on the Ultra2? And which library are you using?

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.

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

Capturing the Diversity of Life - Reorganizing the Protein Space in UniProtKB Advances in genome sequencing technology means that large-scale efforts such as the Earth Biogenome project and the Darwin Tree of Life ...

By the way.. 43% of current TREMBL entries will be dropped soon anyway insideuniprot.blogspot.com/2025/06/capt...

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

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...

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...

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?

aaah guess its "hidden" there :)

Great thanks! Where can I find the one protein per gene option?

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?

💯

Astral Zoom hits >7,000 protein groups & 67,000 precursors — on a 500 SPD EvoSep ENO run.

www.biorxiv.org/content/10.1...

Is that narrow windowed thin-PASEF?

GitHub - pwilmart/quantitative_proteomics_comparison: Comparison of DIA to spectral counting and TMT quantitative techniques using animal lens studies Comparison of DIA to spectral counting and TMT quantitative techniques using animal lens studies - pwilmart/quantitative_proteomics_comparison

DIA, DOA, DUI, DDA, etc. Here is a comparisons of some quantitative proteomics methods from a POV you might not have seen before:
github.com/pwilmart/qua...

Sciex in the game.

Pretty impressed by ScanningSWATH data on the new ZenoTOF 8600.

#TeamMassSpec

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.

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 …

Sure. Why?

..diaPASEF down to 1 ng Hela looks ok (using E.Coli entrapment)

Assessment of false discovery rate control in tandem mass spectrometry analysis using entrapment - Nature Methods A theoretical foundation for entrapment methods is presented, along with a method that enables more accurate evaluation of false discovery rate (FDR) control in proteomics mass spectrometry analysis p...

#TeamMassSpec, Any opinions on why not generally adding the relatively small yeast proteome to the anyway large human search space (*.fasta) as an internal FDR quality control?

www.nature.com/articles/s41...

The videos from the 8th Single-Cell Proteomics Conference (#SCP2025) will be joining this growing YouTube playlist.

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

Aurora Elite™ CSI 15x75 C18 UHPLC column - IonOpticks 15 cm x 75 μm C18 UHPLC packed emitter column, compatible with all Bruker CaptiveSpray sources.

Aurora Elite CSI 15cmx75um. Unfortunately EvoSep methods cannot be modified..

Hi #TeamMassSpec #EvoSep –
Anyone using WhisperZoom for standard inputs (~500 ng)?
Getting great data on timsTOF Pro Ultra (with ICC2), but repeatedly hit overpressure on Aurora columns - forcing me to discard them.
Anyone else seeing this?

A Solid-Phase Extraction Capture (SPEC) workflow in nanoliter volumes for fast, robust and ultrasensitive proteomics Despite great progress, sample preparation remains an area for improvement in proteomics, particularly for low-input samples where conventional protocols lead to losses and incomplete digestion. We pr...

www.biorxiv.org/content/10.1...

SPEC :The better SP3?

Is that related to timePlex?

Introducing the new timsMetabo! A metabolomics focused timsTOF:
- enhanced ion capacity of the dual-stage TIMS-MX ion funnel
- Athen Ion Processor-equipped timsMetabo, up to 300 Hz PRM
Also QSee software (a nightmare at talks, QC or Qsee?).
As found by Biswapriya Misra
www.bruker.com/en/news-and-...