Premium
An improved quantitative mass spectrometry analysis of tumor specific mutant proteins at high sensitivity
Author(s) -
RuppenCañás Isabel,
LópezCasas Pedro P.,
García Fernando,
XiménezEmbún Pilar,
Muñoz Manuel,
Morelli M. Pia,
Real Francisco X.,
Serna Antonio,
Hidalgo Manuel,
Ashman Keith
Publication year - 2012
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201100611
Subject(s) - mass spectrometry , mutant , proteomics , quantitative proteomics , cancer , selected reaction monitoring , computational biology , chemistry , biology , microbiology and biotechnology , tandem mass spectrometry , cancer research , chromatography , biochemistry , gene , genetics
New disease specific biomarkers, especially for cancer, are urgently needed to improve individual diagnosis, prognosis, and treatment selection, that is, for personalized medicine. Genetic mutations that affect protein function drive cancer. Therefore, the detection of such mutations represents a source of cancer specific biomarkers. Here we confirm the implementation of the mutant protein specific immuno‐ SRM (where SRM is selective reaction monitoring) mass spectrometry method of RAS proteins reported by Wang et al. [ Proc. Natl. Acad. Sci. USA 2011, 108, 2444–2449], which exploits an antibody to simultaneously capture the different forms of the target protein and the resolving power and sensitivity of LC ‐ MS / MS and improve the technique by using a more sensitive mass spectrometer. The mutant form G 12 D was quantified by SRM on a QTRAP 5500 mass spectrometer and the MIDAS workflow was used to confirm the sequence of the targeted peptides. This assay has been applied to quantify wild type and mutant RAS proteins in patient tumors, xenografted human tissue, and benign human epidermal tumors at high sensitivity. The limit of detection for the target proteins was as low as 12 amol (0.25 pg). It requires low starting amounts of tissue (ca.15 mg) that could be obtained from a needle aspiration biopsy. The described strategy could find application in the clinical arena and be applied to the study of expression of protein variants in disease.