Here, we present an automated plasma protein digestion method for mass spectrometry-based quantitative proteomic analysis. In this protocol, the liquid transferring and incubation steps for protein denaturation, reduction, alkylation, and trypsin digestion reactions are streamlined and automated. It takes approximately five hours to prepare a 96-well plate with desired precision.
Total time
~5 hours per 96-well plate
Steps
1
Review automated plasma protein digestion principles
Understand the rationale for automating liquid transfer and incubation steps in plasma proteomic sample preparation. Learn how automation improves precision and throughput for mass spectrometry-based quantification.
▶ 00:04
2
Execute automated protein preparation workflow
Operate the automated workstation to perform sequential liquid transfers and incubations for protein denaturation, reduction, alkylation, and enzymatic trypsin digestion on plasma samples in a 96-well plate format.
▶ 01:11
3
Analyze digested peptides by LC-MSMS
Load prepared samples onto liquid chromatography-tandem mass spectrometry instrumentation for peptide separation and quantitative mass spectrometric analysis of the proteome.
▶ 03:28
4
Interpret representative mass spectrometric results
Review sample mass spectra, peptide identifications, and protein quantification data obtained from the automated workflow to validate sample preparation quality and analytical performance.
▶ 04:38