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Screening protein isoforms predictive for cancer using immunoaffinity capture and fast LC‐MS in PRM mode
Author(s) -
Lesur Antoine,
Ancheva Lina,
Kim Yeoun Jin,
Berchem Guy,
Oostrum Jan,
Domon Bruno
Publication year - 2015
Publication title -
proteomics – clinical applications
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.948
H-Index - 54
eISSN - 1862-8354
pISSN - 1862-8346
DOI - 10.1002/prca.201400158
Subject(s) - gene isoform , chemistry , computational biology , chromatography , biology , biochemistry , gene
Purpose We report an immunocapture strategy to extract proteins known to harbor driver mutations for a defined cancer type before the simultaneous assessment of their mutational status by MS. Such a method bypasses the sensitivity and selectivity issues encountered during the analysis of unfractionated complex biological samples. Experimental design Fast LC separations using short nanobore columns hyphenated with a high‐resolution quadrupole‐orbitrap mass spectrometer have been devised to take advantage of fast MS cycle times in conjunction with sharp chromatographic peak widths to accelerate the sample analysis throughput. Such an analytical platform is well suited to analyze simple protein mixtures obtained after immunoaffinity enrichment. Results After establishing the technical performance of the platform, the method was applied to the quantitative profiling of cellular Ras and EGFR protein isoforms, as well as serum amyloid A isoforms in plasma. Conclusions and clinical relevance Immunoaffinity purification combined with fast LC‐MS detection for the detection of driver mutations in tissue and tumor biomarkers in plasma samples can assist clinicians to select an optimal therapeutic intervention for patients.