Bone marrow-derived mesenchymal stem cells (MSCs) lack cell-specific markers that distinguish them from other cell populations such as fibroblasts, adipocytes, and osteogenic cells. Objective: To characterize structural and nanomechanical properties of rat MSCs using atomic force microscopy (AFM). Methods: After dissection of the tibio-femoral bone complex from euthanized 6-week-old male Sprague-Dawley rats, whole bone marrow plugs were obtained, dispersed, and finally cells were suspended Dulbecco's Modified Eagle's Medium-Low Glucose (DMEM-LG) supplemented with 10% fetal bovine serum (FBS) and 1% antibiotic-antimycotic solution. Cells were plated at a density of 5×10⁷ cells/100 mm tissue culture dish and incubated in 95% air/5% CO₂ at 37°C. After 14 days, cells were trypsinized, counted and adjusted to 6×10⁵ cells/ml. A 100 µl aliquot was plated on round glass coverslips (12 mm) that were pretreated with 1% poly-D-lysine and cells were allowed to attach for several hrs before adding medium to the tissue culture dish. On day 4 after plating, MSC/coverslips were mounted on magnetic disks for contact-mode-AFM imaging. During imaging, MSCs were maintained in serum-free DMEM-LG that formed a meniscus between cell surface and the AFM probe in a fluid chamber without O-ring seal. AFM cantilevers had a nominal force constant of k=0.12 N/m and oxide-sharpened Si₃ N₄ tips. Scan sizes varied up to 100×100 mm. Results: The mean surface roughness of MSCs was in the range of 2.87 to 11.4 nm in the Z dimension, reflecting structural variation of the cell surface. The Young's moduli of MSC cell membrane derived from the Hertz model were in the range of 52.3±0.84 kPa to 87.94±0.60 kPa. Conclusion: These structural and nanomechanical properties of MSCs may be potentially useful in the characterization and distinction of bone marrow-derived MSCs from other cell populations.

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