ESF 158細胞

細胞類型： 其他細胞類型

是否是腫瘤細胞： 0

物種來源： 小鼠

品系： ESF 158

ATCC Number： SCRC-1016?

年限： embryo

數(shù)量： 大量

器官來源： 胚胎

生長狀態(tài)： 貼壁生長

運輸方式： 凍存運輸

細胞形態(tài)： 球形

Designations: ESF 158

Depositors: F.Brook

ESF 158細胞Biosafety Level: 1

Shipped: frozen

Medium & Serum: See Propagation

Growth Properties: Adherent

Organism: Mus musculus deposited as mouse

Morphology: spherical colony

Source: Strain: ESF 158

Organ: embryo

Cell Type: embryonic stem cell

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Applications: Coat-color chimera production was high using c2J blastocysts while FVB blastocysts produced a low number of chimeras [PubMed: 11730008].

The ES cells were shown to populate the germ line of two host blastocyst donors, FVB/NJ (FVB) and the coisogenic strain C57BL/6-Tyrc-2J (c2J).

ESF 158細胞The clonal embryonic stem cell line #693 ES ESF 158 was derived from a strain ESF 158 mouse blastocyst [PubMed: 11730008].

Age: embryo

Comments: The clonal embryonic stem cell line #693 ES ESF 158 was derived from a strain ESF 158 mouse blastocyst [PubMed: 11730008]. The ES cells were shown to populate the germ line of two host blastocyst donors, FVB/NJ (FVB) and the coisogenic strain C57BL/6-Tyrc-2J (c2J). Coat-color chimera production was high using c2J blastocysts while FVB blastocysts produced a low number of chimeras [PubMed: 11730008].

Propagation: ATCC complete growth medium: Mouse ES Cell Basal Medium

Atmosphere: air, 95%; carbon dioxide (CO2), 5%

Temperature: 37.0°C

Subculturing: Protocol: Establishing and maintaining your culture: To insure the highest level of viability, be sure to warm media to 37?C before using it on the cells.

Plate mitotically arrested MEF (CF-1) (ATCC SCRC-1040) as a feeder layer at approximately 1.5 to 2.0 X 106 cells/T25 at least one day before plating ESF-158 cells (see product sheet for mitotically arrested MEF for protocol). One hour before thawing the vial of ESF-158 cells, perform a 100% medium change using 10 ml of complete growth medium for ES cells.

Thaw the vial by gentle agitation in a 37°C water bath. To reduce the possibility of contamination, keep the O-ring and cap out of the water. Thawing should be rapid (approximately 90 seconds).

Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by dipping in or spraying with 70% ethanol. All of the operations from this point on should be carried out under strict aseptic conditions.

Transfer the vial's contents plus 5 ml of complete growth medium for ES cells to a 15 ml centrifuge tube. Use an additional 1 ml of media to rinse the vial and transfer the liquid to the 15 ml tube. Add 4 ml of complete growth medium for ES cells to bring the total volume to 10 ml.

Spin the cells at 270 xg for 5 min. ESF 158細胞Aspirate the supernatant and resuspend the pellet in 5 ml of complete growth medium for ES cells.

Add the 5 ml of cell suspension to the T75 flask containing feeder cells and 10 ml complete growth medium for ES cells.

Incubate the culture at 37°C in a humidified 5% CO2/95% air incubator. Perform a 100% medium change every day, passage cells every 1 to 2 days.

Subculturing Procedure: To insure the highest level of viability, be sure to warm media and Trypsin/EDTA to 37°C before using it on the cells. Volumes used in this protocol are for T75 flasks. Proportionally adjust the volumes for culture vessels of other sizes. A split ratio of 1:4 to 1:7 every 1 to 2 days is recommended. Plating densities should range from 3 to 4 X 106 cells/ T75. Note: If the colonies are close to or touching each other the culture is overgrown . Overgrowth will result in differentiation.

Prepare enough flasks with MEFs as stated above in step #1.

Aspirate the medium from the flask(s) with R1 ES cells.

Wash with PBS (Ca+2/Mg+2-free, ATCC cat# SCRR-2201).

Add 3.0 ml of 0.25% (w/v) Trypsin / 0.53 mM EDTA solution (ATCC cat # 30-2101) and place in incubator. After about one minute the ES colonies will dissociate and all cells will detach from the flask.

Dislodge the cells by gently tapping the side of the flask then wash the cells off with 7-10 ml of fresh culture medium. Triturate cells several times with a 10 ml pipette in order to dissociate the cells into a single-cell suspension.

Spin the cells at 270 xg for 5 min. Aspirate the supernatant.

Resuspend in 30 to 50 ml of fresh culture medium, depending on the split ratio.

Aspirate the medium from 4 to 7 feeder layer flasks and replace it with 15 ml/flask of R1 cell suspension.

Incubate the culture at 37°C in a humidified 5% CO2/95% air incubator. Perform a 100% medium change every day, passage cells every 1 to 2 days.

Interval: Every one to two days

Preservation: Freeze medium: ESF 158細胞Complete growth medium supplemented with an additional 10% FBS and 10% DMSO.

Storage temperature: liquid nitrogen vapor phase

Related Products: recommended serum: ATCC SCRR-30-2020

References: 16173597: Brook FA, et al. The derivation of highly germline-competent embryonic stem cells containing NOD-derived genome. Diabetes. 52:205-208, 2003. PubMed: 12502514

16173598: Brook FA, Gardner RL. The origin and efficient derivation of embryonic stem cells in the mouse. Proc. Natl. Acad. Sci. USA. 94: 5709-5712, 1997. PubMed: 9159137