By Kursad Turksen
Now in volumes, this thoroughly up-to-date and improved version of Embryonic Stem Cells: equipment and Protocols presents a various selection of simply reproducible mobile and molecular protocols for the manipulation of nonhuman embryonic stem cells. quantity one, Embryonic Stem mobilephone Protocols: Isolation and Characterization, moment version, presents a various number of easily reproducible mobile and molecular protocols for the isolation, upkeep, and characterization of embryonic stem cells. the second one quantity, Embryonic Stem mobile Protocols: Differentiation versions, moment version, covers cutting-edge tools for deriving many sorts of differentiating cells from ES cells. A significant other CD presents digital colour types of all illustrations within the publication. jointly, the 2 volumes light up for either newcomers and specialists our present knowing of the biology of embryonic stem cells and their application in general tissue homeostasis and regenerative drugs functions.
Read or Download Embryonic Stem Cell Protocols 2nd Edition, Volume 1: Isolation And Characterization (Methods in Molecular Biology Vol 329) PDF
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Extra resources for Embryonic Stem Cell Protocols 2nd Edition, Volume 1: Isolation And Characterization (Methods in Molecular Biology Vol 329)
Sample text
Derivation of ES Cell Cultures Cell cultures can only be initiated from permissive medaka strains such as HB32C (2,11). The blastoderm, the cell mass of an MBE, consists of 1000 cells opposite to the oil droplets (Fig. 1B). Freshly seeded MBEs are 30–40 µm in size, irregular in shape, and active in amoebalike movement by forming pseudopodia (Fig. 1C). They divide quickly and begin to attach 12–18 h after seeding at 28°C. An even distribution and a proper cell density are important to prevent differentiation into neural and fibroblastlike cells.
Thaw nucleotide stock at 37°C before use. 15. Fertilized, freshly laid chicken eggs. 16. Lipopolysaccharide (LPS) from E. coli serotype O127:B8 (100 mg; Sigma, cat. no. L3880). 17. Penicillin-streptomycin 100X (100 mL; Gibco BRL, cat. no. 15140-122). 18. 100 × 20-mm tissue culture dish (Becton Dickinson, cat. no. 353003). 19. 15-mL centrifuge tube (Sumilon, Tokyo, Japan; cat. no. MS-56150). 20. Filter paper rings (see Note 5). 21. Glass or plastic tube (see Note 6). 1. Media 1. Tissue culture medium for IN24 cell line: IN24 cells (chicken monocytic leukemia cells) (10) are maintained in IMDM supplemented with 10% heat-inactivated FBS and 1% penicillinstreptomycin (10% FBS-IMDM).
No. 161-0363). 4. 2% Tween-20, 3% bovine serum albumin. 4. 5. Anti-STAT3 (749-769) rabbit antibody (1 mg/mL; Calbiochem, San Diego, CA; cat. no. 57609). 6. Anti-Phospho-STAT3 (Tyr705) antibody (100 µL; Cell Signaling Technology, Beverly, MA; cat. no. 9131). 7. Horseradish peroxidase-labeled antirabbit IgG (1 mg/mL; KPL, Gaithersburg, MD; cat. no. 474-1516). 8. ECL plus Western blotting detection reagents (Amersham Pharmacia Biotech, cat. no. RPN2132). 3. 1. 1. Isogen RNA Extraction Chicken LIF cDNA was cloned using mRNA from LPS-stimulated IN24 cells, and chicken LIF mRNA was most abundant in adult chicken liver and thymus (9).