Oligodendrocyte progenitor cell compositions
An oligodendrocyte and composition technology, which is applied in the field of cell biology and can solve the problems of complex, expensive, non-scalable custom reagents, etc.
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[0127] One aspect of the present disclosure includes a container comprising a composition, wherein the composition comprises a population of cells comprising a plurality of oligodendrocyte progenitor cells (OPCs), and wherein the population of cells comprises less than 15% of the undesired cell type. In one aspect, undesired cell types include cells of epithelial lineage. In one aspect, cells of epithelial lineage are characterized by the presence of one or more markers selected from the group consisting of K7 and PCK. In one aspect, the population of cells comprises less than 2% K7 positive cells. In one aspect, the population of cells comprises less than 0.2% K7 positive cells. In certain aspects, the population of cells comprises less than 5% K7 positive cells. In one aspect, populations of cells share a common genetic background. In one aspect, the population of cells has not undergone cell enrichment. In one aspect, the container is configured for cryopreservation. ...
Embodiment 1
[0138] Example 1 - Method for preparing OPC populations including pretreatment
[0139] Undifferentiated human embryonic stem cells (uhESC) from the H1 line (WA01; Thomson JA, Itskovitz-EldorJ, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM. Embryonic stem cell lines derived from human blastocysts (Embryonic stem cell lines derived from human blastocysts). S Science. 1998 Nov 6; 282(5391): 1145-7) in KO-DMEM diluted 1:30 GFR with X-VIVO 10 supplemented with 80ng / mL bFGF (ThermoFisher, PHG0263) and 0.5ng / mL TGF-β1 (R&D System, cat# 240-B) cultured on culture medium. Daily medium changes were complete starting 2 days after subculturing the cells. Undifferentiated hESCs were subcultured using collagenase and manual scraping.
[0140] Expose uhESCs to a cocktail of small molecules to boost cells prior to induction of differentiation. In subculture passages before initiation of differentiation, cells were incubated at approximately 0.8-1.0 × 10 5 cells / cm 2 Inoc...
Embodiment 2
[0144] Example 2 - Comparison of Alternative Methods for Preparing OPC Populations Including Pretreatment
[0145] Undifferentiated human embryonic stem cells (uhESC) from the H1 line (WA01; Thomson JA, Itskovitz-EldorJ, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM. Embryonic stem cell lines derived from human blastocysts (Embryonic stem cell lines derived from human blastocysts). Science. 1998 Nov 6; 282(5391): 1145-7) on one of the following matrices: 1:30 dilution in KO-DMEM GFR, recombinant laminin 521 (Corning self-coat cat. no. 354221 or PureCoat), or vitronectin (StemCell Technologies, cat. no. 07180) in cell adhesion dilution buffer (StemCell Technologies, cat. No. 07183) with X- VIVO 10 Medium. Daily medium changes were complete starting 2 days after subculturing the cells. Use collagenase and manual scraping or other non-enzymatic means such as 0.5 mM EDTA in PBS (Life Technologies, cat. no. 15575-020) or ReLeSR TM (Stem Cell Technologies, Cat. No...
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