Internal calibrants allow high accuracy peptide matching between MALDI imaging MS and LC-MS/MS

Area of Science:

• Biomolecular analysis
• Mass spectrometry
• Proteomics

Background:

• Unambiguous identification of analytes in MALDI imaging mass spectrometry (MALDI-IMS) is a significant challenge.
• Matching MALDI-IMS data with LC-MS/MS identified peptides is hindered by the variability in MALDI-TOF/TOF MS measurements.
• Current external calibration methods limit mass accuracy in MALDI-IMS, complicating data correlation with LC-MS/MS results.

Purpose of the Study:

• To develop and validate a method for internal calibration of tryptic peptide MALDI-IMS data.
• To improve the mass accuracy of MALDI-IMS measurements for formalin-fixed paraffin-embedded ovarian cancer tissues.
• To enhance the matching of MALDI-IMS m/z values with LC-MS/MS identified peptide m/z values.

Main Methods:

• Applied internal calibration to MALDI-IMS datasets from tryptic peptides of ovarian cancer tissue sections.
• Utilized internal peak features within the spectra for calibration.
• Confirmed improved m/z accuracy by matching MALDI-IMS data with LC-MS/MS identified peptides and performing subsequent in situ MS/MS analysis.

Main Results:

• Internal calibration significantly reduced the m/z error when matching MALDI-IMS data to LC-MS/MS identified peptides.
• The improved mass accuracy facilitated more reliable correlation between MALDI-IMS and LC-MS/MS datasets.
• Follow-up MS/MS analysis confirmed the accuracy of the matched peptide identifications.

Conclusions:

• Internal calibration is a crucial method for enhancing the mass accuracy of tryptic peptide MALDI-IMS.
• This approach overcomes limitations of external calibration, enabling more precise analyte identification.
• Internal calibrants should be routinely incorporated into tryptic peptide MALDI-IMS experiments for improved data quality and comparability.