A potentially scalable method for the harvesting of hMSCs from microcarriers
journal contributionposted on 16.06.2014, 13:05 authored by Alvin W. Nienow, Qasim Rafiq, Karen CoopmanKaren Coopman, Christopher Hewitt
The use of hMSCs for allogeneic therapies requiring lot sizes of billions of cells will necessitate large-scale culture techniques such as the expansion of cells on microcarriers in bioreactors. Whilst much research investigating hMSC culture on microcarriers has focused on growth, much less involves their harvesting for passaging or as a step towards cryopreservation and storage. A successful new harvesting method has recently been outlined for cells grown on SoloHill microcarriers in a 5 L bioreactor . Here, this new method is set out in detail, harvesting being defined as a two-step process involving cell ‘detachment’ from the microcarriers’ surface followed by the ‘separation’ of the two entities. The new detachment method is based on theoretical concepts originally developed for secondary nucleation due to agitation. Based on this theory, it is suggested that a short period (here 7 min) of intense agitation in the presence of a suitable enzyme should detach the cells from the relatively large microcarriers. In addition, once detached, the cells should not be damaged because they are smaller than the Kolmogorov microscale. Detachment was then successfully achieved for hMSCs from two different donors using microcarrier/cell suspensions up to 100 mL in a spinner flask. In both cases, harvesting was completed by separating cells from microcarriers using a Steriflip® vacuum filter. The overall harvesting efficiency was >95% and after harvesting, the cells maintained all the attributes expected of hMSC cells. The underlying theoretical concepts suggest that the method is scalable and this aspect is discussed too.
The authors would like to acknowledge the Biotechnology and Biological Sciences Research Council (BBSRC; UK) Bioprocessing Research Industries Club (BRIC), the Engineering and Physical Sciences Research Council (EPSRC; UK) [grant number EP/F500491/1] and Lonza GmbH (Cologne, Germany) for their support and funding.
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