Method of deriving progenitor cell line
a progenitor cell and stem cell technology, applied in the field of cell biology, molecular biology and genetics, can solve the problems of heterogeneous cell populations, inefficient protocols, and inability to generate other cell types that may have a deleterious effect on tissue repair and regeneration, and achieve the effect of promoting or retarding the self-renewal or differentiation of stem cells
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Benefits of technology
Problems solved by technology
Method used
Image
Examples
example 1
Methods: Derivation of E-RoSH Cell Lines
[0204]Embryonic stem cells (ESCs) are induced to differentiate to form embryoid bodies (Ebs) using the methycellulose-based approach described in Lim et al, Blood. 1997; 90:1291-1299)
[0205]Day 3 to day 6 embryoid bodies are harvested, dissociated into single cell suspensions by collagenase digestion (Robertson E J. Embryo-derived stem cell lines. In: Robertson E J, ed. Teratocarcinomas and embryonic stem cells: a practical approach. Oxford: IRL Press Limited; 1987:71-112) and plated on at a density of 1-5×105 cells per 10 cm feeder plate. After about a week, the cells proliferated and differentiated into a complex mixture of cell types.
[0206]Colonies of rapidly dividing cells resembling embryo-derived RoSH cells are picked and expanded sequentially to a 48-well plate, 24-well plate, 6-well plate and then a 10 cm plate. The culture from each colony is named E-RoSH1, 2, 3 . . . in the sequence in which each culture is established.
[0207]Each of t...
example 2
Methods: Derivation of HuES9.E Mesenchymal Stem Cell (MSC)-Like Cell Lines
[0209]HuES9 cells are cultured as previously described in Cowan et al, N Engl J Med. 2004; 350:1353-1356.
[0210]To derive HuES9.E MSC-like cells, HuES9 cells are split 1:4 onto gelatinized feeder-free plates in using HuES9 culture media. Confluent cultures are typsinized and split 1:4. Differentiation into adipocytes, and osteocytes is performed as previously described (Barberi et al, PLoS Med. 2005; 2:e161).
[0211]BM MSCs are prepared as previously described in Pittenger et al, Science. 1999; 284:143-147.
[0212]Genomic PCR for mouse- and human-specific repeat sequences are performed as previously described in Que et al, In Vitro Cell Dev Biol Anim. 2004; 40:143-149.
example 3
Methods: RT-PCR Analysis
[0213]Total RNA is prepared using standard protocols and are quantified using, respectively, the RiboGreen RNA Quantification kit and the PicoGreen dsDNA Quantification kit (Molecular Probes, Eugene, Oreg.).
[0214]Quantitative RT-PCR is performed using TaqMan® primers (Applied Biosystems, Foster City, Calif.).
PUM
| Property | Measurement | Unit |
|---|---|---|
| thickness | aaaaa | aaaaa |
| concentration | aaaaa | aaaaa |
| diameter | aaaaa | aaaaa |
Abstract
Description
Claims
Application Information
Login to View More