Comparative LC-MS/MS analysis of optimal cutting temperature (OCT) compound removal for the study of mammalian proteomes.
Primary tissue samples are valuable resources for investigators interested in understanding disease. In order to maximize the information content that can be gained from these precious samples, proper storage, handling, and preparation are essential. Some tissue preservation techniques utilize the cryopreservation medium, optimal cutting temperature (OCT) compound. While this medium provides benefits for traditional molecular studies, certain components can interfere with mass spectrometric analyses. Mass spectrometry based proteomics is a growing field with many applications for disease research. Our goal is to determine a reliable method for separating the proteins from the contaminating species in OCT embedded samples, thus making these samples compatible with mass spectrometric analyses. The novel applications of ether-methanol precipitation, filter-aided sample preparation (FASP), and SDS-PAGE provide researchers with protocols for removing OCT contaminating species from valuable samples. The results presented in this study show that all three methods reproducibly remove OCT; however, precipitation and FASP outperform SDS-PAGE by common proteomic metrics.
Department of Chemistry and Biochemistry, University of Notre Dame, 251 Nieuwland Science Hall, Notre Dame, IN 46556, USA. ahummon@nd.edu.
Leigh A
LA
Weston
Amanda B
AB
Hummon
D000818:Animals; D002417:Cattle; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D020543:Proteome / Q000032:analysis*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013696:Temperature
D020543:Proteome