Mesenchymal stem cells (MSCs) have various characteristic properties such as self-renewal and multilineage differentiation capability, making them promising tools to be used in clinics in therapeutic settings as a cell-based therapy. For MSCs to be used for therapeutic applications, their prolonged expansion in culture is required to provide a sufficient number of cells. During the expansion in culture, MSCs experience only a restricted number of population doublings, and then they enter senescence. Becoming senescent impairs MSCs’ proliferation capability, colony-forming efficacy, and ability to differentiate into various lineages and these limit their application in clinics. Thus, monitoring the senescence and aging of MSCs is essential for their use in therapeutic applications. In this chapter, we summarized the protocols to assess the aging of MSCs through senescence-associated β-galactosidase and colony-forming unit assay.