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A label‐free mass spectrometry method for relative quantitation of β‐tubulin isotype expression in human tumor tissue
Author(s) -
Miller Leah M.,
Huang Yang ChiaPing,
Xiao Hui,
Isaac Sylvie,
Sève Pascal,
Dumontet Charles,
Band Horwitz Susan,
Hogue Angeletti Ruth
Publication year - 2012
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.201200018
Subject(s) - isotype , tubulin , taxane , immunohistochemistry , biology , microbiology and biotechnology , antibody , microtubule , immunology , cancer , monoclonal antibody , genetics , breast cancer
Purpose Quantitation of β‐tubulin isotype expression in taxane resistant human tumor tissue has been difficult to achieve because of the limited availability of validated antibodies. Here we present a label‐free MS method to quantitate relative expression levels of β‐tubulin isotypes. Experimental design Using isotype‐specific reporter peptides, we determined relative β‐tubulin isotype expression levels in human lung tumor tissue. Results Four reporter peptides were chosen to quantitate the βI/βII, βIV, βIII, and βV tubulin isotypes. These peptides were validated using human cancer cell lines. The label‐free method was then used to determine β‐tubulin isotype expression in nine human lung tumor samples, which had been described as high or low βIII‐tubulin expressing using immunohistochemistry. It was found that βI/βII (accounting for 18.7–65.7% of total β‐tubulin) and βIVa/βIVb (26.3–79.1%) were the most abundant isotypes and that the βIII (0–8.9%) and βV (1.0–10.4%) were less abundant in the tissue. We also categorized the samples as high or low βIII‐tubulin expressing. Conclusion and clinical relevance With this method we can determine the relative expression levels of β‐tubulin isotypes in human tumor tissue. This method will facilitate studies assessing the use of tubulin isotypes as biomarkers of taxane resistance.