Isolation, enumeration, and expansion of human mesenchymal stem cells in culture

Human bone marrow (BM) contains a population of non-hematopoietic stem cells also termed stromal cells, mesenchymal cells or multipotent mesenchymal stromal cells (MSCs). These cells have unique stem cell-like properties including their ability to self-renew, differentiate into multiple tissue types, and modulate immune cell responses through paracrine effects. These properties have positioned mesenchymal cells as biological agents in clinical trials for various diseases since the 1990s. Mesenchymal cells have been isolated from various tissues and cultured using various media and methods resulting in a lack of standardization in culture methods for these cells. Consequently, cells cultured in different laboratories exhibit different characteristics of MSC-like cells. This chapter outlines protocols for optimal isolation, enumeration, and expansion of human MSCs from BM in fetal bovine serum (FBS)-containing medium, as well as in xeno-free medium.