📋 Workflow
Full workflow, products add-to-cart directly1
Protein digestion
4–16 hReduce/alkylate then digest protein into peptides with trypsin
💡 Tip Trypsin cleaves after K/R
⚠️ Caution Thorough reduction/alkylation avoids disulfide interference
🧪 Materials for this step (click to shop by spec/brand)
2
Desalting/purification
30 minRemove salts/detergents and concentrate peptides with C18 reverse-phase column or StageTip
💡 Tip Desalting improves ionization efficiency
⚠️ Caution Elute fully to avoid peptide loss
🧪 Materials for this step (click to shop by spec/brand)
3
LC separation and ionization
2–4 hSeparate peptides by nano-LC gradient, ionize by electrospray into the mass spectrometer
💡 Tip Longer gradients boost resolution and identifications
⚠️ Caution Stable LC conditions for reproducibility
🧪 Materials for this step (click to shop by spec/brand)
4
Data acquisition and analysis
2–4 hAcquire MS/MS spectra, search against protein database with software and quantify
💡 Tip Set FDR to control false positives
⚠️ Caution Calibrate for mass accuracy
🧪 Materials for this step (click to shop by spec/brand)
⚠️ 5 Common Beginner Mistakes
- ① Incomplete digestion reduces coverage
- ② Salt/detergent contamination suppresses ionization
- ③ Calibration drift gives mass error
- ④ Incomplete database lowers identification
- ⑤ Improper FDR yields false positives
❓ FAQ
+Can MS identify PTMs?
Yes, it can localize phosphorylation, acetylation, glycosylation and other PTM sites.
+How to increase identifications?
Optimize gradient, enrich low-abundance, add separation dimensions or higher resolution.
+What determines sensitivity?
Ionization efficiency, sample amount, instrument sensitivity and desalting purity.