Abstract 3207: Application of an antibody-free, highly sensitive PRISM-SRM proteomics approach for monitoring low abundance proteins and protein isoforms in cell lines and tumor tissue.

Abstract

Abstract Stable-isotope dilution coupled to targeted mass spectrometry, applying selected reaction monitoring (SRM), provides good selectivity, reproducibility, and sensitivity rendering it highly suitable for robust and high-throughput protein quantification. Historically, SRM sensitivity for measuring protein concentration in biofluids such as blood plasma has been limited to the low μg/mL level. Application of front-end sample preparation methods such as immunoaffinity depletion and strong cation exchange (SCX) chromatographic fractionation has improved the SRM detection limit to low ng/mL level. The ability to detect and quantify low abundance proteins present in blood plasma/serum at sub-ng/mL levels, however, relies on the availability of specific affinity reagents (e.g., antibodies that have the associated disadvantage of an expensive, lengthy, and high failure rate de novo process). To address this limitation, we have recently developed an antibody-free, targeted quantification capability that enables protein quantification of prostate-specific antigen and other exogenous proteins at 50-100 pg/mL levels in plasma/serum. Unlike the traditional SCX fractionation approach where all resulting fractions are analyzed by the second dimension LC-SRM, this new capability, PRISM (high-pressure high-resolution separations with intelligent selection and multiplexing), uses high resolution. Citation Format: Jintang He, Xuefei Sun, Tujin Shi, Athena Schepmoes, Thomas Fillmore, Vladislav Petyuk, Fang Xie, Marina Gritsenko, Feng Yang, Naoki Kitabayashi, Sung Suk Chae, Mark Rubin, Javed Siddiqui, John Wei, Arul Chinnaiyan, Wei-Jun Qian, Richard Smith, Jacob Kagan, Sudhir Srivastava, Tao Liu, Karin Rodland, David Camp. Application of an antibody-free, highly sensitive PRISM-SRM proteomics approach for monitoring low abundance proteins and protein isoforms in cell lines and tumor tissue. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3207. doi:10.1158/1538-7445.AM2013-3207