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In this chapter, we describe methods in which proteins can be extracted for proteomic analysis using a shotgun approach. The chapter outlines important in vitro techniques and data analysis that can be applied to investigate the proteome dynamics.Cell-mediated cytotoxicity plays an important role in several fundamental immunological processes and is crucial for biological evaluation in in vitro studies. In order to determine the immunological activities of the cells, an assay should be safe, reproducible, and cost-effective. Here, we p