Mesenchymal stem cells (MSCs) are an attractive target for clinical study as therapeutic agents. However, current multilayer flatbed culture-expansion paradigms are cumbersome, time consuming, and typically limited in the ability to monitor cell characteristics during the growth process.
We have used the Mobius® CellReady 3-L bioreactor in combination with microcarriers and human mesenchymal stem cells (hMSC) as a model. Following expansion, we have comprehensively compared the characteristics of cells grown in the bioreactor with those grown in standard two-dimensional (2D) cultures. It has been demonstrated that MSCs grown on microcarriers in a stirred-tank bioreactor showed the same phenotypic and genetic markers as cells grown in standard 2D cultures:
Adherent stem cells (hMSCs) can be successfully expanded in a 3-L bioreactor with microcarriers
hMSCs expanded in bioreactor showed no difference in flow-cytometry profiles, differentiation abilities, or functional assays from cells produced in 2D CellFactory system.
The 3L CellReady bioreactor process for hMSC can be controlled and monitored to achieve consistent cell production.
Millipore’s Mobius® CellReady 3-L bioreactor is a single-use, stirred-tank bioreactor ideal for cell culture process development. Arriving preassembled and gamma irradiated, the CellReady 3-L bioreactor significantly reduces the assembly time, downtime, and maintenance time associated with traditional glass bioreactors. To ensure maximum operational flexibility, the CellReady 3-L bioreactor comes standard with prefitted weldable tubing lines, two available sparging options, a vent line, and integrated side sampling, addition, and drain ports. Designed to replace traditional bench-scale glass bioreactors, the CellReady 3-L bioreactor is compatible with most standard controller configurations.Materials and Methods
MSCs (EMD Millipore SCR108) were cultured under static conditions with low-glucose DMEM (Invitrogen® 11054), 10% FBS (HyClone® SH30070.03), Pen/Strep (EMD Millipore TMS-AB2-C), L-Glutamine (EMD Millipore TMS-002-C), and 8 ng/mL bFGF Human Recombinant (EMD Millipore GF003AF-MG) in T-150 flasks coated with gelatin (EMD Millipore ES-006B). Low-oxygen conditions were used for 2D propagation as well as the attachment phase in which the MSCs were attached to the collagen-coated microcarriers (Solohill C102-1521) in Petri dishes.Summary
Clinical demand for MSCs is driving the need for development of robust large-scale production. Expansion of hMSCs in stirred-tank bioreactors using microcarriers is the first step to establishing a scalable, single-use, and efficient production process. The 3-L CellReady bioreactor presents several advantages over multilayer flatbed cultures, including improved ease of use, reduced medium consumption, and the ability to monitor process parameters and cell health.
We have demonstrated the ability to expand mesenchymal stem cells microcarriers in a single-use, stirred-tank bioreactor. hMSCs expanded in that bioreactor showed no difference in flow cytometry profiles, differentiation abilities, or functional assays compared with cells produced in a 2D CellFactory system. The 3-L CellReady bioreactor process for hMSC can be easily controlled and monitored to achieve consistent cell production and is an ideal platform for large-scale production.
Robert Shaw is commercial director of the stem cell initiative at EMD Millipore Corp., 80 Ashby Road, Bedford, MA 01730; 1-781-533-2511, mobile 1-978-761-0758.