An optimized MALDI MSI protocol for spatial detection of tryptic peptides in fresh frozen prostate tissue

MALDI MS imaging (MSI) is a powerful analytical tool for spatial peptide detection in heterogeneous tissues. Proper sample preparation is crucial to achieve high quality, reproducible measurements. Here we developed an optimized protocol for spatially resolved proteolytic peptide detection with MALD...

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Bibliographic Details
Published in:Proteomics (Weinheim) 2022-05, Vol.22 (10), p.e2100223-n/a
Main Authors: Høiem, Therese S., Andersen, Maria K., Martin‐Lorenzo, Marta, Longuespée, Rémi, Claes, Britt S.R., Nordborg, Anna, Dewez, Frédéric, Balluff, Benjamin, Giampà, Marco, Sharma, Animesh, Hagen, Lars, Heeren, Ron M.A., Bathen, Tone F., Giskeødegård, Guro F., Krossa, Sebastian, Tessem, May‐Britt
Format: Article
Language:English
Subjects:
Biomarkers
Biopolymer denaturation
Digestion
fresh frozen
Humans
Localization
MALDI mass spectrometry imaging
Male
Peptides
Prostate
Prostate - metabolism
Prostate cancer
prostate tissue
Protein denaturation
Proteins
Proteolysis
proteomics
Quality assessment
Sample preparation
Spatial discrimination
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods
Tissues
Trypsin
Trypsin - metabolism
Tryptic peptides
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Summary:MALDI MS imaging (MSI) is a powerful analytical tool for spatial peptide detection in heterogeneous tissues. Proper sample preparation is crucial to achieve high quality, reproducible measurements. Here we developed an optimized protocol for spatially resolved proteolytic peptide detection with MALDI time‐of‐flight MSI of fresh frozen prostate tissue sections. The parameters tested included four different tissue washes, four methods of protein denaturation, four methods of trypsin digestion (different trypsin densities, sprayers, and incubation times), and five matrix deposition methods (different sprayers, settings, and matrix concentrations). Evaluation criteria were the number of detected and excluded peaks, percentage of high mass peaks, signal‐to‐noise ratio, spatial localization, and average intensities of identified peptides, all of which were integrated into a weighted quality evaluation scoring system. Based on these scores, the optimized protocol included an ice‐cold EtOH+H2O wash, a 5 min heating step at 95°C, tryptic digestion incubated for 17h at 37°C and CHCA matrix deposited at a final amount of 1.8 μg/mm2. Including a heat‐induced protein denaturation step after tissue wash is a new methodological approach that could be useful also for other tissue types. This optimized protocol for spatial peptide detection using MALDI MSI facilitates future biomarker discovery in prostate cancer and may be useful in studies of other tissue types.
ISSN:1615-9853
1615-9861
1615-9861
DOI:10.1002/pmic.202100223