Fig. 1
From: Functional fingerprinting of human mesenchymal stem cells using high-throughput RNAi screening
Workflow for high-throughput RNAi screening experiments in primary human MSCs. a Bone marrow is aspirated from healthy human donors and following density gradient centrifugation the mononuclear cell fraction is seeded into culture flasks. Initial MSC colonies are separated by their plastic adherence and expanded. MSCs are further characterized by immunophenotyping using fluorescence flow cytometry and their ability to differentiate towards adipogenic and osteogenic lineages. For high-throughput screening, MSCs were used at passage 4 and reverse transfected in cell culture plates. b Optimization of siRNA transfection conditions for assay time (at 200 cells/well) (b) and varying cell numbers (at 72 h) (c). Cell viability was determined using a CellTiterGlo assay. Ctrl, pGL3; UBC, siRNA against Ubiquitin C