ATCC® Number: CRL-2922™ Price:

Designations:

EA.hy926

Depositors: CS Edgell

Biosafety Level: 1

Shipped: frozen

Medium & Serum: See Propagation

Growth Properties: adherent

Organism: Homo sapiens （human）

Morphology: endothelial

CRL-2922 EA.hy926 人臍靜脈細胞融合細胞

Source: Tissue: somatic cell hybrid

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Restrictions: For-profit institutions must obtain license from University of North Carolina, Chapel Hill.

Antigen Expression: Factor VIII-related antigen; Homo sapiens, expressed

DNA Profile （STR）: Amelogenin: X

CSF1PO: 10,11,12

D13S317: 11

D16S539: 11,12

D5S818: 11

D7S820: 8,9,10

THO1: 6,8,9.3

TPOX: 8,9

vWA: 14,17

CRL-2922 EA.hy926 人臍靜脈細胞融合細胞

Comments: The human umbilical vein cell line, EA.hy926, was established by fusing primary human umbilical vein cells with a thioguanine-resistant clone of A549 by exposure to polyethylene glycol （PEG）. Hybrid clones were selected in HAT medium and screened for factor VIII-related antigen. EA.hy926 cells have been maintained for more than 100 population doublings （PDLs）. Electron photomicrographs demonstrate cytoplasmic distribution of Weibel-Palade bodies and tissue-specific organelles, characteristics of differentiated endothelial cell functions such as angiogenesis, homeostasis/thrombosis, blood pressure and inflammation. [PubMed: 6407019] [PubMed: 2079463]

Propagation: ATCC complete growth medium: The base medium for this cell line is ATCC-formulated Dulbeccos Modified Eagles Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10.

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

Temperature: 37.0°C

Subculturing: Protocol: Volumes used in this protocol are for 75 sq cm flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.

1.Remove and discard culture medium.

2.Briefly rinse the cell layer with Ca++/Mg++ free Dulbeccos phosphate-buffered saline （D-PBS） or 0.25 （w/v） Trypsin - 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.

3.Add 2.0 to 3.0 ml of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed （usually within 5 to 15 minutes）.

CRL-2922 EA.hy926

Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37C to facilitate dispersal.

4.Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.

5.Transfer cell suspension to a centrifuge tube and spin at approximay 125 X g for 5 to 10 minutes. Discard supernatant.

6.Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 2 X 10（3） to 3 X 10（3） viable cells/sq. cm is recommended.

7.Incubate cultures at 37C. Subculture when cell concentration reaches between 8 X 10（4） and 1 X 10（5） cells/sq. cm.

Interval: Twice a week

Subc*tion Ratio: A subc*tion ratio of 1:6 to 1:10 is recommended

Medium Renewal: Every 2 to 3 days

Preservation: Freeze medium: Complete Growth medium, 95; DMSO, 5

Storage temperature: liquid nitrogen vapor phase

Related Products: Recommended medium （without the additional supplements or serum described under ATCC Medium）:ATCC 30-2002

recommended serum:ATCC 30-2020

CRL-2922 EA.hy926

0.25 （w/v） Trypsin - 0.53 mM EDTA in Hank BSS （w/o Ca++, Mg++）:ATCC 30-2101

phosphate-buffered saline:ATCC 30-2200

Cell culture tested DMSO:ATCC 4-X

Erythrosin B vital stain solution:ATCC 30-2404

Trypan Blue vital stain solution:ATCC 30-2402

References: 92902: Edgell CJ, et al. Permanent cell line expressing human factor VIII-related antigen established by hybridization. Proc. Natl. Acad. Sci. USA 80: 3734-3737, 1983. PubMed: 6407019

92903: Bauer J, et al. In vitro model of angiogenesis using a human endothelium-derived permanent cell line: contributions of induced gene expression, G-proteins, and integrins. J. Cell. Physiol. 153: 437-449, 1992. PubMed: 1280276

92904: Edgell CJ, et al. Endothelium specific Weibel-Palade bodies in a continuous human cell line, EA.hy926. In Vitro Cell. Dev. Biol. 26: 1167-1172, 1990. PubMed: 2079463

CRL-2922 EA.hy926

92905: Rieber AJ, et al. Extent of differentiated gene expression in the human endothelium-derived EA.hy926 cell line. Thromb. Haemostasis 69: 476-480, 1993. PubMed: 8322270