HUV-EC-C [HUVEC] (ATCC^® CRL-1730^™)

The base medium for this cell line is F-12K Medium (ATCC 30-2004). To make the complete growth medium, add the following components to 440 mL of the base medium:

• 5 mL of a 10 mg/mL stock heparin solution (prepared from Sigma catalog #H3393) for a final concentration of 0.1 mg/mL heparin in complete growth medium 
  • Dissolve 1 g Heparin in 100 mL basal F-12K  and filter to make a 10 mg/mL stock solution
• 5 mL endothelial cell growth supplement (ECGS; BD Biosciences catalog # 354006)
  • Rehydrate 1 vial using 5 mL basal F-12K 
• 50 mL fetal bovine serum (FBS; ATCC 30-2020)

Note: A high quality ECGS prepared from bovine neural tissue (BD Biosciences catalog # 354006 or equivalent) should be used to propagate CRL-1730. It is best to initiate the cells with the highest recommended concentration of ECGS. Moderate to heavy debris and numerous floating cells may be routinely observed in cultures of HUV-EC-C cells.  Retain the floating cells by gentle centrifugation and add back to the adherent population.

CRL-1730 is a slow growing cell line that has a roughly estimated doubling time of 5 to 6 days. Cultures should be fully fluid changed every 48 hours. The cells should only be allowed to go 72 hours without fluid changing when the density is less than 50% confluent. Perform full fluid changes rather than media additions.

This cell line produces a lot of floaters and debris especially at higher densities. Cells detach before completely filling in to 100% confluence. It is recommended to subculture the cells when 80 to 90% confluent to avoid excessive floaters. Floating cells are viable and if pronounced, they should be spun down and reseeded back into the growing culture.

1. Remove and discard culture medium.
2. Briefly rinse the cell layer with 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).
   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 37°C to facilitate dispersal.
4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
5. Add appropriate aliquots of the cell suspension to new culture vessels.
6. Incubate cultures at 37°C.

Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:3 is recommended

Medium Renewal: Two to three times per week

Seeding Density: 8.0 x 10³ to 3.0 x 10⁴ viable cells/cm²

Molestina RE, et al. Characterization of a strain of Chlamydia pneumoniae isolated from a coronary atheroma by analysis of the omp1 gene and biological activity in human endothelial cells. Infect. Immun. 66: 1370-1376, 1998. PubMed: 9529055

Zahedi K. Characterization of the binding of serum amyloid P to laminin. J. Biol. Chem. 272: 2143-2148, 1997. PubMed: 8999915

Lindstrom AL, et al. An in vitro model for toxin-mediated vascular leak syndrome: ricin toxin A chain increases the permeability of human endothelial cell monolayers. Blood 90: 2323-2334, 1997. PubMed: 9310483

Soker S, et al. Inhibition of vascular endothelial growth factor (VEGF)-induced endothelial cell proliferation by a peptide corresponding to the exon 7-encoded domain VGEF165. J. Biol. Chem. 272: 31582-31588, 1997. PubMed: 9395496

Li Y, et al. Mast cell granules potentiate endotoxin-induced interleukin-6 production by endothelial cells. J. Leukocyte Biol. 62: 211-216, 1997. PubMed: 9261335

Soker S, et al. Characterization of novel vascular endothelial growth factor (VEGF) receptors on tumor cells that bind VEGF165 via its exon 7-endoded domain. J. Biol. Chem. 271: 5761-5767, 1996. PubMed: 8621443

Hoshi H, McKeehan WL. Brain- and liver cell-derived factors are required for growth of human endothelial cells in serum-free culture. Proc. Natl. Acad. Sci. USA 81: 6413-6417, 1984. PubMed: 6333682

1. CRL-1730 morphology and culture condition
   

   
   Date Updated: 2/21/2014

2. Number of ATCC® CRL-1730 cells in a confluent flask
   

   
   Date Updated: 2/5/2013

3. ATCC® CRL-1730 vs. ATCC® CRL-2873
   

   
   Date Updated: 2/4/2013

Molestina RE, et al. Characterization of a strain of Chlamydia pneumoniae isolated from a coronary atheroma by analysis of the omp1 gene and biological activity in human endothelial cells. Infect. Immun. 66: 1370-1376, 1998. PubMed: 9529055

Zahedi K. Characterization of the binding of serum amyloid P to laminin. J. Biol. Chem. 272: 2143-2148, 1997. PubMed: 8999915

Lindstrom AL, et al. An in vitro model for toxin-mediated vascular leak syndrome: ricin toxin A chain increases the permeability of human endothelial cell monolayers. Blood 90: 2323-2334, 1997. PubMed: 9310483

Soker S, et al. Inhibition of vascular endothelial growth factor (VEGF)-induced endothelial cell proliferation by a peptide corresponding to the exon 7-encoded domain VGEF165. J. Biol. Chem. 272: 31582-31588, 1997. PubMed: 9395496

Li Y, et al. Mast cell granules potentiate endotoxin-induced interleukin-6 production by endothelial cells. J. Leukocyte Biol. 62: 211-216, 1997. PubMed: 9261335

Soker S, et al. Characterization of novel vascular endothelial growth factor (VEGF) receptors on tumor cells that bind VEGF165 via its exon 7-endoded domain. J. Biol. Chem. 271: 5761-5767, 1996. PubMed: 8621443

Hoshi H, McKeehan WL. Brain- and liver cell-derived factors are required for growth of human endothelial cells in serum-free culture. Proc. Natl. Acad. Sci. USA 81: 6413-6417, 1984. PubMed: 6333682