Optimization of a serum-free growth medium. (a) Fluorescence-based quantification of human MSC proliferation in deficient serum-free medium (SFM) supplemented with various combinations of recombinant human growth factors or in serum-containing medium (SCM) by using the CyQUANT NF Cell Proliferation Assay. D, growth factor-deficient SFM; P, SFM supplemented with recombinant human PDGF-BB; B, SFM supplemented with recombinant human bFGF; T, SFM supplemented with recombinant human TGF-β1. (b) Morphology of passage 4 human MSCs expanded in (a) SCM and (b) SFM. Magnification, 10× objective. (c) Recovered four-donor pooled human MSCs (passage 5) expanded under optimized conditions for eight additional passages (48 days) in SFM or SCM.