Abstract
One of the most significant difficulties in studying clinical specimens such as cancer stem cells by mass spectrometry based proteomics approaches arises from the small sample size which limits the use of conventional proteome profiling procedures. The ideal strategy to avoid sample loss is to entirely diminish sample preparation, but current mass spectrometry techniques do not allow direct identification/quantification of proteins present in complex biological matrices. In this chapter, we describe a workflow that combines a single-tube cell lysis procedure using PPS silent surfactant (3-[3-(1,1-bisalkyloxyethyl)pyridin-1-yl]propane-1-sulfonate) and a capillary isoelectric focusing (CIEF) step to specifically analyze a small number of cells. This workflow can be directly coupled to reversed-phase liquid chromatography (RPLC) followed by nanoflow rate electrospray ionization mass spectrometry (nanoESI- MS) to study the global proteome expression pattern with the proteins from 104 pancreatic cancer tissue stem cells (equal to <1 μg of protein material) which is 100 times less than the number of cells needed in a typical proteome profiling experiment.
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Dai, L., Li, C., Lubman, D.M. (2011). Development of Micro-scale Sample Preparation and Prefractionation Methods in LC-MS-Based Proteomic Studies. In: Ivanov, A., Lazarev, A. (eds) Sample Preparation in Biological Mass Spectrometry. Springer, Dordrecht. https://doi.org/10.1007/978-94-007-0828-0_44
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DOI: https://doi.org/10.1007/978-94-007-0828-0_44
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