Abstract
Rapidly evolving cell-based therapies towards clinical trials demand alternative approaches for efficient expansion of adherent cell types such as human mesenchymal stem cells (hMSCs). Using microcarriers (100–300 µm) in a stirred tank bioreactor offers considerably enhanced surface to volume ratio of culture environment. However, downstream purification of the harvested cell product needs to be addressed carefully due to distinctive features and fragility of these cell products. This work demonstrates a novel alternative approach which utilizes inertial focusing to separate microcarriers (MCs) from the final cell suspension. First, we systematically investigated MC focusing dynamics inside scaled-up curved channels with trapezoidal and rectangular cross-sections. A trapezoidal spiral channel with ultra-low-slope (Tan(α) = 0.0375) was found to contribute to strong MC focusing (~300 < Re < ~400) while managing high MC volume fractions up to ~1.68%. Accordingly, the high-throughput trapezoidal spiral channel successfully separated MCs from hMSC suspension with total cell yield~94% (after two passes) at a high volumetric flow rate of ~30 mL/min (Re~326.5).
Highlights
Off-the-shelf therapies transplanting human mesenchymal stem cells, derived mainly from bone-marrow, adipose tissue, and umbilical cord blood tissue[1], are widely adopted due to hMSCs’ regenerative, immunosuppressive, and multipotent features[2,3]
The results show that the trapezoidal spiral enhances a positive secondary flow, which contributes to driving particles to the inner wall, and alters the distribution of secondary flow
Because inertial focusing of MCs near the inner wall cannot be interpreted solely as a result of positive secondary flow without considering the shear force; we investigated MC focusing dynamics experimentally due to the lack of a shear-gradient force model exclusively for spiral channels
Summary
Off-the-shelf (allogeneic) therapies transplanting human mesenchymal stem cells (hMSCs), derived mainly from bone-marrow, adipose tissue, and umbilical cord blood tissue[1], are widely adopted due to hMSCs’ regenerative, immunosuppressive, and multipotent features[2,3]. Among different adherent cell bioreactors, employing suspension scaffolds so-called microcarriers (MCs), ~100–300 μm in diameter, within a stirred tank has been widely recognized[7,13]; recently it was demonstrated within a 50-L bioreactor that a 43-fold expansion of hMSCs could be reached in 11 days[14]. We report on the advancement of an alternative method using inertial focusing – shown recently to be scalable for filtration of large-scale lot size in the order of liter per min[17,18,19,20]. Removal of microcarriers from hMSCs suspension was accomplished by inertial focusing with ~99% purity while cell harvest yield reached ~94%
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