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Supplemental Protocol: Yeast Lysis with PIXUL Multi-Sample Sonicator

Lyse 96 Saccharomyces cerevisiae cultures in parallel and feed straight into bottom-up LC-MS/MS — VIB Proteomics Core validated end-to-end workflow, no bead-beating, no probe sonication.

This 25-step protocol takes you from yeast cell pellet to LC-MS-ready peptides in a single high-throughput workflow: PIXUL megasonic lysis in 5% SDS / 50 mM TEAB (Pulse N=50, PRF=1 kHz, Burst Rate=20 Hz, 30 or 60 min process time), DTT reduction, iodoacetamide alkylation, S-Trap 96-well cleanup, overnight trypsin digestion at 37 C, and a three-step peptide elution into MS vials. Developed and validated by the VIB Proteomics Core at the Vlaams Instituut voor Biotechnologie (Belgium). Written for yeast proteomics scientists, mass spec core facility staff, and bench scientists working with S. cerevisiae overnight cultures.

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  • VIB Proteomics Core validated workflow for S. cerevisiae lysis and bottom-up proteomics sample prep — institutional endorsement from one of Europe's premier life sciences research institutes.
  • PIXUL yeast lysis parameters in 96-well round bottom plates (Cat. No. 53139): 5% SDS / 50 mM TEAB (pH 7-8) at 100 uL per well, Pulse N=50, PRF=1 kHz, Burst Rate=20 Hz, Process Time 30 or 60 min — disrupts the rigid chitin-glucan-mannan cell wall without bead-beating or probe sonication.
  • Complete 25-step bottom-up workflow from yeast pellet through BCA quantification, DTT/IAA reduction-alkylation, S-Trap acid-and-methanol cleanup, 1:100 overnight trypsin digestion, and three-step peptide elution (aqueous TEAB, 0.2% formic acid, 50% ACN/0.2% FA) into SpeedVac-dried MS vials.
  • Built-in A/B comparison pathway (Steps 6-8) for benchmarking PIXUL against the traditional gold standard of Lysing Matrix Y bead-beating on a Qiagen TissueLyser plus probe sonication, with agarose-gel aliquot checkpoints for direct lysis-efficiency comparison.
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