RWPE2-W99

CRL-2853 ^™

Product category: Human cells
Organism: Homo sapiens, human
Cell type: epithelial cell
Morphology: epithelial
Tissue: Prostate
Disease: Normal
Applications: 3D cell culture
Product format: Frozen
Storage conditions: Vapor phase of liquid nitrogen

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Documentation

Product sheet

BSL 2

ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.

Cells contain Human papillomavirus type 18 (HPV-18) sequences

ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.

General

Specific applications: The RWPE2-W99 cell line was derived, in 1999, from the RWPE-2 (ATCC CRL-11610) cell line by cloning in soft agar to select cells that show high expression of Ki-ras.

Characteristics

Growth properties: Adherent
Derivation: The RWPE2-W99 cell line was derived, in 1999, from the RWPE-2 (ATCC CRL-11610) cell line by cloning in soft agar to select cells that show high expression of Ki-ras. RWPE2-W99 cells show strong expression of Ki-ras and have characteristics similar to those of RWPE-2 cells. RWPE-2 cells were derived from RWPE-1 cells (ATCC CRL-11609) by transformation with Ki-ras using the Kirsten murine sarcoma virus (Ki-MuSV) to establish the RWPE-2 cells (PubMed: 9214605). Epithelial cells from the peripheral zone of a histologically normal adult human prostate were transfected with a plasmid carrying one copy of the human papilloma virus 18 (HPV-18) genome to establish the RWPE-1 cell line (ATCC CRL-11609). A family of tumorigenic cell lines that mimics multiple steps in prostate cancer progression was also derived from RWPE-1 cells by exposure to N-methyl-N-nitrosourea (MNU): WPE1-NA22 (ATCC CRL-2849), WPE1-NB14 (ATCC CRL-2850), WPE1-NB11 (ATCC CRL-2851) and WPE1-NB26 (ATCC CRL-2852).The depositor reports that this cell line was screened for Hepatitis B and C, and human immunodeficiency viruses, and was found to be negative.
Age: 54 years
Ethnicity: White
Gender: Male
Karyotype: The depositor reports that at passage 44, a majority of the cells had a near diploid chromosome number of 48; X, -Y.
Tumorigenic: Yes;
Yes, in nude mice
(RWPE2-W99 cells form small tumors when injected subcutaneously, with Matrigel, in nude mice.)(Mukta M Webber, personal communication)
Yes, the cells form colonies in soft agar
(small colonies are formed)(Mukta M Webber, personal communication)
Antigen expression: Kallikrein 3, KLK3 (prostate-specific antigen, PSA); Homo sapiens, expressed (upon exposure to androgen)
(upon exposure to androgen)
Genes expressed: cytokeratin 8; cytokeratin 18
Expression markers: Androgen receptor, expressed
Comments: The RWPE2-W99 cell line was derived, in 1999, from the RWPE-2 (ATCC CRL-11610) cell line by cloning in soft agar to select cells that show high expression of Ki-ras. RWPE2-W99 cells show strong expression of Ki-ras and have characteristics similar to those of RWPE-2 cells. RWPE-2 cells were derived from RWPE-1 cells (ATCC CRL-11609) by transformation with Ki-ras using the Kirsten murine sarcoma virus (Ki-MuSV) to establish the RWPE-2 cells (PubMed: 9214605). Epithelial cells from the peripheral zone of a histologically normal adult human prostate were transfected with a plasmid carrying one copy of the human papilloma virus 18 (HPV-18) genome to establish the RWPE-1 cell line (ATCC CRL-11609). A family of tumorigenic cell lines that mimics multiple steps in prostate cancer progression was also derived from RWPE-1 cells by exposure to N-methyl-N-nitrosourea (MNU): WPE1-NA22 (ATCC CRL-2849), WPE1-NB14 (ATCC CRL-2850), WPE1-NB11 (ATCC CRL-2851) and WPE1-NB26 (ATCC CRL-2852).
The depositor reports that this cell line was screened for Hepatitis B and C, and human immunodeficiency viruses, and was found to be negative.

Handling information

Unpacking and storage instructions

Check all containers for leakage or breakage.
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.

Complete medium

The base medium for this cell line is provided by Invitrogen (GIBCO) as part of a kit: Keratinocyte Serum Free Medium (K-SFM), Kit Catalog Number 17005-042. This kit is supplied with each of the two additives required to grow this cell line (bovine pituitary extract (BPE) and human recombinant epidermal growth factor (EGF). To make the complete growth medium, you will need to add the following components to the base medium:

0.05 mg/ml BPE - provided with the K-SFM kit

5 ng/ml EGF - provided with the K-SFM kit. NOTE: Do not filter complete medium.

Temperature

37°C

Atmosphere

95% Air, 5% CO₂

Handling procedure

Handling Procedure for Frozen Cells

To insure the highest level of viability, thaw the vial and initiate the culture as soon as possible upon receipt. If upon arrival, continued storage of the frozen culture is necessary, it should be stored in liquid nitrogen vapor phase and not at -70°C. Storage at -70°C will result in loss of viability.

SAFETY PRECAUTION: ATCC highly recommends that protective gloves and clothing always be used and a full face mask always be worn when handling frozen vials. It is important to note that some vials leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vessel exploding or blowing off its cap with dangerous force creating flying debris.

1. 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 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 to7 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 37°C in a suitable incubator. A 5% CO₂ in air atmosphere is recommended if using the medium described on this product.

Subculturing procedure

Protocol:

Remove and discard culture medium.
Briefly rinse the cell layer with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS).
Add 2.0 to 3.0 ml (to a T-25 flask) or 3.0 to 4.0 ml (to a T-75 flask) of 0.05% Trypsin - 0.53 mM EDTA solution, diluted 1:1 with D-PBS, and place flask in a 37C incubator for 5 to 8 minutes. Observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 10 minutes).
Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach.
Add 6.0 to 8.0 ml of 0.1% Soybean Trypsin Inhibitor (or 2% fetal bovine serum in D-PBS), as appropriate, and aspirate cells by gently pipetting.
Transfer cell suspension to centrifuge tube and spin at approximately 125 x g for 5 to 7 minutes.
Discard supernatant and resuspend cells in fresh serum-free growth medium. Add appropriate aliquots of cell suspension to new culture vessels. An inoculum between 2 X 10(4) to 4 X 10(4) viable cells/sq. cm is recommended.
Incubate cultures at 37C. We recommend that you maintain cultures at a cell concentration between 4 X 10(4) and 8 X 10(4) cells/sq. cm.

Cells grown under serum-free or reduced serum conditions may not attach strongly during the 24 hours after subculture and should be disturbed as little as possible during that period.

Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:5 is recommended

Medium Renewal: Every 48 hours

Note: Subculture cells before they reach confluence. Do not allow cells to become confluent.

Quality control specifications

Mycoplasma contamination: Not detected
Population doubling time: Approximately 36 hrs
STR profiling: Amelogenin: X,Y
CSF1PO: 13
D13S317: 8,14
D16S539: 9,11
D5S818: 12,15
D7S820: 10,11
TH01: 8,9.3
TPOX: 8,11
vWA: 14,18
D3S1358: 15,16
D21S11: 29,31
D18S51: 14,16
Penta_E: 5,12
Penta_D: 10,13
D8S1179: 10,14
FGA: 24,25
D19S433: 13
D2S1338: 17,20

History

Depositors: MM Webber
Year of origin: 1999

Legal disclaimers

Intended use

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.

Warranty

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.

Disclaimers

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.

Permits & Restrictions

Import Permit for the State of Hawaii

If shipping to the U.S. state of Hawaii, you must provide either an import permit or documentation stating that an import permit is not required. We cannot ship this item until we receive this documentation. Contact the Hawaii Department of Agriculture (HDOA), Plant Industry Division, Plant Quarantine Branch to determine if an import permit is required.

MORE INFORMATION ABOUT PERMITS AND RESTRICTIONS

References

Curated Citations

Bello D, et al. Androgen responsive adult human prostatic epithelial cell lines immortalized by human papillomavirus 18. Carcinogenesis 18: 1215-1223, 1997. PubMed: 9214605

Webber MM, et al. Acinar differentiation by non-malignant immortalized human prostatic epithelial cells and its loss by malignant cells. Carcinogenesis 18: 1225-1231, 1997. PubMed: 9214606

Webber MM, et al. Prostate specific antigen and androgen receptor induction and characterization of an immortalized adult human prostatic epithelial cell line. Carcinogenesis 17: 1641-1646, 1996. PubMed: 8761420

Okamoto M, et al. Interleukin-6 and epidermal growth factor promote anchorage-independent growth of immortalized human prostatic epithelial cells treated with N-methyl-N-nitrosourea. Prostate 35: 255-262, 1998. PubMed: 9609548

Webber MM, et al. Immortalized and tumorigenic adult human prostatic epithelial cell lines: characteristics and applications. Part I. Cell markers and immortalized nontumorigenic cell lines. Prostate 29: 386-394, 1996. PubMed: 8977636

View All Curated Citations for this Product

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