Proteomics refers to the analysis of expression, localization, functions, posttranslational modifications, and interactions of proteins expressed by a genome at a specific condition and at a specific time. Mass spectrometry (MS)-based proteomic methods have emerged as a key technology for unbiased systematic and high-throughput identification and quantification of complex protein mixtures. These methods have the potential to reveal unknown and novel changes in protein interactions and assemblies that regulate cellular and physiological processes. Both gel-based (one-dimensional [1D] gel electrophoresis, two-dimensional [2D] polyacrylamide gel electrophoresis, 2D difference in-gel electrophoresis [DIGE]) and gel-free (liquid chromatography [LC], capillary electrophoresis) approaches have been developed and utilized in a variety of combinations to separate proteins prior to mass spectrometric analysis. Detailed protocols for global proteomic analysis from adipose-derived stem cells (ASCs) using two central strategies, 2D-DIGE-MS and 2D-LC-MS, are presented here.