iTRAQ™ labeling coupled with LC-MALDI mass spectrometry for monitoring temporal response of colorectal cancer cells to butyrate treatment.

Abstract

Mass spectrometry (MS)-based quantitative proteomics plays important roles in drug discovery. In this chapter, we describe a stable isotope labeling technique which employs 4-plex iTRAQ(™) isobaric reagents coupled with two-dimensional (2-D) liquid chromatography (LC) and MALDI-TOF/TOF MS, for a temporal study of HCT-116 colon carcinoma cells treated with butyrate. Butyrate is a short-chain fatty acid fermentation by-product of fiber that can induce temporal cell maturation, from the early phase of growth arrest, to differentiation, and to the activation of apoptotic cascades. Our quantitative proteomics study uncovered several integrated cellular processes and pathways involved in growth arrest, apoptosis, and metastasis. Selected protein targets are validated by real-time PCR and western blotting.