Systematic profiling of signaling pathways on the proteome level can reveal functionally important proteins that can be used in precision diagnostics, prediction of treatment response and development of personalized therapies. Mass spectrometry (MS)-based proteomics has become the method of choice in this area, given the ability to study thousands of proteins simultaneously. This chapter covers the basics and current state of bottom-up proteomics workflow, including sample preparation, liquid chromatography separations, MS instrumentation, data acquisition strategies, quantification techniques, and data analysis. Moreover, we describe the recent applications of proteomics in the precision medicine field.