Springer Protocols: Abstract: Isolation and Culture of Bone Marrow-Derived Human Multipotent Stromal Cells (hMSCs)

Isolation and Culture of Bone Marrow-Derived Human Multipotent Stromal Cells (hMSCs)

By: Margaret Wolfe³, Radhika Pochampally⁴, William Swaney³, Roxanne L. Reger³

Abstract

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We have developed protocols whereby a total of 30–90 × 10⁶ hMSCs with an average viability greater than 90% can be produced in a single multilevel Cell Factory from a relatively small (1–3 mL) bone marrow aspirate in 14–20 d. It is possible to generate as many as 5 × 10⁸ multipotent stromal cells (MSCs) from a single sample, depending on the number of Cell Factories seeded from the initial isolated hMSCs. Briefly, mononuclear cells are collected from a bone marrow aspirate by density gradient centrifugation. The cells are cultured overnight and the adherent cells are allowed to attach to the flask. Nonadherent cells are removed and the culture expanded for 7–10 d with periodic feeding of the cells. The cells are then harvested and seeded at low density (60–100 cells/cm²) into Nunc Cell Factories. The cells are allowed to expand for an additional 7–10 d, and are then harvested.

Affiliation(s): (3) Center for Gene Therapy, Tulane University Health Sciences Center, New Orleans, LA
(4) Center for Gene Therapy and Department of Pharmacology, Tulane University Health Sciences Center, New Orleans, LA

Book Title: Mesenchymal Stem Cells: Methods and Protocols

Series: Methods in Molecular Biology | Volume: 449 | Pub. Date: Apr-01-2008 | Page Range: 3-25 | DOI: 10.1007/978-1-60327-169-1_1

Subject: Cell Biology

Key Words: Mesenchymal stem cells - MSCs - isolation - culture - multipotent

References

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