RPE-J

RPE-J is a retinal pigment epithelial (RPE) cell line derived from primary cultures of RPE cells taken from 7-day-old Long-Evans rats.

The RPE-J clone was selected for an epithelioid morphology and for expression of circumferential staining with antibody against the tight junction protein 1, (Tjp1, ZO-1).

The cells express a transformed phenotype at the permissive temperature (33°C), and a non-transformed phenotype at the non-permissive temperature (40°C). They must be cultured at the permissive temperature and do not grow at 37°C.

When RPE-J cells are grown on nitrocellulose filters coated with a thin layer of Matrigel in the presence of 10(-8) M retinoic acid for 6 days at 33°C and then switched to the non-permissive temperature of 40°C for 33 to 36 hours, they acquire a differentiated polarized RPE phenotype. Under these conditions, RPE-J cells exhibit circumferential staining for the tight-junction protein ZO-1 and acquire a transepithelial resistance of 350 ohms/cm2. RefNabi IR, et al. Immortalization of polarized rat retinal pigment epithelium. J. Cell Sci. 104: 37-49, 1993. PubMed: 8383696

RPE-J is the only established RPE cell line that maintains epithelial cell surface polarity. The cells retain many properties of RPE including expression of the rat RPE marker RET-PE2 and the ability to phagocytose latex beads.RefNabi IR, et al. Immortalization of polarized rat retinal pigment epithelium. J. Cell Sci. 104: 37-49, 1993. PubMed: 8383696

A culture submitted to the ATCC in July 1995 was found to be contaminated with Mycoplasma hyorhinis and was cured by a 21-day treatment with BM Cycline. The cells were assayed for mycoplasma by the Hoechst stain and the standard culture test over a six-week period following treatment and all tests were negative.

1. Check all containers for leakage or breakage.
2. Remove the frozen cells from the dry ice packaging and immediately place the cells at a temperature below ­-130°C, preferably in liquid nitrogen vapor, until ready for use.

Dulbecco's modified Eagle's medium with 4 mM L-glutamine adjusted to contain 1.5g/L sodium bicarbonate and 4.5 g/L glucose and supplemented with 0.1 mM non-essential amino acids (Gibco 11140-050), 96%; fetal bovine serum, 4%

1. Thaw the vial by gentle agitation in a 33°C water bath. To reduce the possibility of contamination, keep the O-ring and cap out of the water.  Thawing should be rapid (approximately 2 minutes).
2. 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.
3. Transfer the vial contents to a centrifuge tube containing  9.0 mL complete culture medium and spin at approximately 125 x g for 5 to 7 minutes.
4. Resuspend cell pellet with the recommended complete medium (see the specific batch information for the culture recommended dilution ratio) and dispense into a 25 cm² or a 75 cm² culture flask.  It is important to avoid excessive alkalinity of the medium during recovery of the cells.  It is suggested that, prior to the addition of the vial contents, the culture vessel containing the complete growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH (7.0 to 7.6).
5. Incubate the culture at 33°C in a suitable incubator.  A 5% CO₂ in air atmosphere is recommended if using the medium described on this product sheet.

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 that 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. Transfer cell suspension to a centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes. Discard supernatant.
6. Resuspend the cell pellet in fresh growth medium.  Add appropriate aliquots of cell suspension to new culture vessels.
7. Place culture vessels in incubators at 33°C.

The product is provided 'AS IS' and the viability of ATCC^® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid.  Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.

This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use. Any proposed commercial use is prohibited without a license from ATCC.

While ATCC uses reasonable efforts to include accurate and up-to-date information on this product sheet, ATCC makes no warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. ATCC does not warrant that such information has been confirmed to be accurate or complete and the customer bears the sole responsibility of confirming the accuracy and completeness of any such information.

This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.

Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.