Internal calibrants allow high accuracy peptide matching between MALDI imaging MS and LC-MS/MS.
One of the important challenges for MALDI imaging mass spectrometry (MALDI-IMS) is the unambiguous identification of measured analytes. One way to do this is to match tryptic peptide MALDI-IMS m/z values with LC-MS/MS identified m/z values. Matching using current MALDI-TOF/TOF MS instruments is difficult due to the variability of in situ time-of-flight (TOF) m/z measurements. This variability is currently addressed using external calibration, which limits achievable mass accuracy for MALDI-IMS and makes it difficult to match these data to downstream LC-MS/MS results. To overcome this challenge, the work presented here details a method for internally calibrating data sets generated from tryptic peptide MALDI-IMS on formalin-fixed paraffin-embedded sections of ovarian cancer. By calibrating all spectra to internal peak features the m/z error for matches made between MALDI-IMS m/z values and LC-MS/MS identified peptide m/z values was significantly reduced. This improvement was confirmed by follow up matching of LC-MS/MS spectra to in situ MS/MS spectra from the same m/z peak features. The sum of the data presented here indicates that internal calibrants should be a standard component of tryptic peptide MALDI-IMS experiments.
Adelaide Proteomics Centre, School of Molecular and Biomedical Science, The University of Adelaide, Adelaide 5005, Australia.
Johan O R
JOR
Gustafsson
Adelaide Proteomics Centre, School of Molecular and Biomedical Science, The University of Adelaide, Adelaide 5005, Australia.
James S
JS
Eddes
Adelaide Proteomics Centre, School of Molecular and Biomedical Science, The University of Adelaide, Adelaide 5005, Australia.
Stephan
S
Meding
Adelaide Proteomics Centre, School of Molecular and Biomedical Science, The University of Adelaide, Adelaide 5005, Australia.
Tomas
T
Koudelka
Robinson Institute, Research Centre for Reproductive Health, School of Paediatrics and Reproductive Health, The University of Adelaide, Adelaide 5005, Australia.
Martin K
MK
Oehler
Chemokine Biology Laboratory, School of Molecular and Biomedical Science, The University of Adelaide, Adelaide 5005, Australia.
Shaun R
SR
McColl
Adelaide Proteomics Centre, School of Molecular and Biomedical Science, The University of Adelaide, Adelaide 5005, Australia. Electronic address: peter.hoffmann@adelaide.edu.au.
Peter
P
Hoffmann
D000595:Amino Acid Sequence; D002138:Calibration; D002277:Carcinoma / Q000737:chemistry / Q000378:metabolism; D002853:Chromatography, Liquid / Q000379:methods / Q000592:standards; D003952:Diagnostic Imaging / Q000379:methods / Q000592:standards; D005260:Female; D006801:Humans; D008867:Microtomy / Q000379:methods / Q000592:standards; D015588:Observer Variation; D010051:Ovarian Neoplasms / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping / Q000379:methods / Q000592:standards; D010455:Peptides / Q000032:analysis* / Q000737:chemistry; D015203:Reproducibility of Results; D020539:Sequence Analysis, Protein / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods / Q000592:standards*
D010455:Peptides