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RPTEC/TERT1; Renal Proximal Tubules; Human (Homo sapiens) Item ID: ATCCRL4031

Permits and Restrictions

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Organism Homo sapiens, human
Tissue Renal cortex; proximal tubules, epithelium
Cell Type Epithelial cells immortalized with pLXSN-hTERT retroviral transfection
Product Format frozen 1.0 mL
Morphology Epithelial-like
Culture Properties Adherent
Biosafety Level 2  [Cells contain SV40 viral DNA sequences]

Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Age Adult
Gender Male
Applications These cells are proposed to be a valuable model system not only for cell biology, but also toxicology, drug screening, biogerontology and tissue engineering.
Storage Conditions Liquid nitrogen vapor phase
Images
Antigen Expression Antigen expression: This cell line is positive for epithelial marker pan-cytokeratin (immunocytochemistry)(verified at ATCC), positive for epithelial cell adhesion molecule E-cadherin (immunocytochemistry) (verified at ATCC),  The RPTEC/TERT1 cells express both Aminopeptidase N (verified at ATCC) and γ-Glutamyl Transferase (GGT) that are located in the brush border of the renal proximal tubular epithelium.
Comments

The RPTEC/TERT1 cells specifically respond to parathyroid hormone (PTH) but not arginine vasopressin (AVP), and react with enhanced ammonia genesis on lowering of the environmental pH.

The RPTEC/TERT1 cells exhibit sodium-dependent uptake of phosphate as well as intact functionality of the megalin/cubilin transport system.

RPTEC/TERT1 cells show the characteristic morphology and functional properties of normal proximal tubular epithelial cells.

At high cell densities, the RPTEC/TERT1 cells form characteristic "domes", maintain ultrastructural organization with tight junctions, densely packed microvilli and primary cilium, indicating functional cell polarization.

When cultured on the Corning™ Transwell™ Permeable membrane cell culture insert, the RPTEC/TERT1 cells at confluence form intact functional barrier as indicated by stabilized Trans-Epithelial Electrical Resistance (TEER) across the membrane.
Complete Growth Medium

The base medium for this cell line is ATCC-formulated DMEM:F12 Medium (ATCC®30-2006™).

To make the complete growth medium, add hTERT RPTEC Growth Kit (ATCC®ACS-4007™) to the base medium. The final concentration for each growth kit component in the complete hTERT immortalized RPTEC growth medium is as follows:  

  • 5 pM triiodo-L-thyronine
  • 10 ng/mL recombinant human EGF
  • 3.5 µg/mL ascorbic acid
  • 5.0 µg/mL human transferrin
  • 5.0 µg/mL insulin
  • 25 ng/mL prostaglandin E1
  • 25 ng/mL hydrocortisone
  • 8.65 ng/mL sodium selenite
  • 0.1 mg/mL G418
  • 1.2 mg/mL sodium bicarbonate

Note: Do not filter complete medium.

This medium is formulated for use with a 5% CO2 in air atmosphere.

 
Subculturing Volumes are given for a 75 cm2 flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
  1. Subculture when the culture is about 90% confluence. Expected cell yield is between 1.5 x 105 and 2 x 105 viable cells/cm2.
  2. Remove and discard culture medium.
  3. Add 2.0 to 3.0 mL of 0.25% (w/v) trypsin-0.53 mM EDTA solution to the flask and observe cells under an inverted microscope until the cell layer is dispersed (usually within 15 to 20 minutes). Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Place at 37°C to facilitate dispersal.
  4. To stop trypsinization, add 2.0 to 3.0 mL of 0.1% Soybean Trypsin Inhibitor and aspirate cells by gently pipetting.
  5. Transfer cell suspension to a 15-mL centrifuge tube and spin at approximately 250 x g for 5 to 10 minutes.
  6. Discard supernatant and resuspend cells in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 4 to 6 x 104 viable cells/cm2 is recommended.
  7. Incubate cultures at 37°C.
Subcultivation ratio: A subcultivation ratio of 1:3 to 1:4 is recommended. Medium renewal: 2 to 3 times weeklyNote: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 13 in Culture Of Aminal Cells: A Manual of Basic Techniques by R. Ian Freshney, 5th edition, published by Wiley-Liss, N.Y., 2005.
Cryopreservation Freeze medium: DMEM:F12, 90%; DMSO, 10% Storage temperature: liquid nitrogen vapor phase
Culture Conditions Temperature: 37°C Atmosphere: air, 95%; carbon dioxide (CO2), 5%
Volume 1.0 mL
STR Profile CSF1PO: 11 D13S317: 11, 13 D16S539: 11, 12 D5S818: 9, 11 D7S820: 10 THO1: 9, 9.3 TPOX: 8, 11 vWA: 16, 18 Amelogenin: XY
Population Doubling Level (PDL) As part of our quality control, we have tested this cell line for its ability to grow for a minimum of 15 population doublings after recovery from cryopreservation. In addition, it has been verified that no gross changes are observed in karyotype and morphology during the first 10 population doublings.
Name of Depositor R Grillari-Voglauer
Year of Origin July 2004
References

Wieser M, et al. hTERT alone immortalizes epithelial cells of renal proximal tubules without changing their functional characteristics. Am. J. Physiol. Renal Physiol. 295: 1365-1375, 2008. PubMed: 18715936

Bodnar AG, et al. Extension of life-span by introduction of telomerase into normal human cells. Science 279: 349-352, 1998. PubMed: 9454332

Freshney RI. Culture of Animal Cells: A Manual of Basic Technique, 5th edition. New York: Wiley Liss; 2005. For more information on enzymatic dissociation and subculturing of cell lines see Chapter 13.

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