Colony forming unit assays

Abstract

Mesenchymal stem/stromal cells (MSCs) have been extensively investigated for their potential to regenerate tissue, to modulate the immune system, and their wound healing properties in over 350 clinical trials worldwide. MSCs from various tissues such as adipose, bone, and others are currently being studied in clinical trials in indications for ischemic, inflammatory, autoimmune, and degenerative disorders. As a result, numerous isolation protocols have been published. This chapter provides a simple protocol whereby a total of 80-100 million human MSCs, with an average viability greater than 90 %, can be produced from a relatively small (1-3 mL) bone marrow aspirate in 14-20 days using double stack culture chambers. MSCs were originally referred to as fibroblastoid colony forming cells because one of their characteristic features is adherence to tissue culture plastic and generation of colonies when plated at low densities. The efficiency with which they form colonies still remains an important assay for the quality of cell preparations. To assess the quality of cell preparations, two different colony forming unit (CFU) assays are also provided.