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PNEC30

CRL-2930

PNEC30 is a neuroendocrine cell line that was isolated in 2002 from the prostate of a 6-month-old male mouse. The cell line was established from CR2-TAg prostate tumors and metastases. It can be used for genetic and/or pharmacologic studies of the regulation of NE cell proliferation, differentiation, and tumorigenesis.
Product category
Animal cells
Organism
Mus musculus, transgenic, mouse, transgenic
Cell type
neuroendocrine cell
Morphology
neuronal
Tissue
Prostate
Disease
Prostate Cancer; Neuroendocrine
Applications
3D cell culture
Cancer research
Product format
Frozen
Storage conditions
Vapor phase of liquid nitrogen
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Documentation

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.

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.

Required Products

These products are vital for the proper use of this item and have been confirmed as effective in supporting functionality. If you use alternative products, the quality and effectiveness of the item may be affected.

Detailed product information

General

Specific applications
PNEC cell lines should be useful for genetic and/or pharmacologic studies of the regulation of NE cell proliferation, differentiation, and tumorigenesis.

Characteristics

Growth properties
Adherent
Derivation
Prostate neuroendocrine cancer (PNEC) cell lines were established from CR2-TAg prostate tumors and metastases.
Age
6 months
Gender
Male
Tumorigenic
Yes;
tumorigenic in BALB/c mice
Genes expressed
mAsh1 transcription factor
Expression markers
Gamma-aminobutyric acid (GABAA), expressed
Comments

GeneChip analyses of cell lines harvested at different passages, and as xenografted tumors, indicated that PNECs express consistent features ex vivo and in vivo and share a remarkable degree of similarity with primary CR2-TAg prostate neuroendocrine (NE) tumors. 

PNECs express mAsh1 (mouse homolog proneural gene complex achaete-scute), a basic helix-loop-helix (bHLH) transcription factor essential for NE cell differentiation in other tissues. PNEC cell lines should be useful for genetic and/or pharmacologic studies of the regulation of NE cell proliferation, differentiation, and tumorigenesis.

PNEC30 cells, when cultured on non-coated surfaces, grow in suspension as multicellular aggregates that resemble the neurospheres of cultured neural stem cells.

Handling information

Unpacking and storage instructions
  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.
Complete medium
The base medium for this cell line is Neural Progenitor Basal Medium, which is supplied as part of the NPMM Neural Progenitor Maintenance Medium Bullet Kit available from Lonza/Clonetics Inc., Catalog No. CC-3209. To make the complete growth medium, add the following components to 500 ml of the base medium:
  • additives that are supplied with the kit (ATCC does not use gentamycin-amphotericin B)
  • heat-inactivated fetal bovine serum (FBS) to a final concentration of 10%
  • bovine pituitary extract (BPE) (Lonza/Clonetics, Inc., Catalog No. CC-4009) to a final concentration of 0.3%

Note: Do not filter complete medium.
Temperature
37°C
Atmosphere
95% Air, 5% CO2
Handling procedure

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.


Note: These cells are cultured on poly-D-lysine coated vessels (BD BioCoat® Cellware, BD Biosciences, Cat. No. 356524 for 75 cm2 flasks) which are additionally coated with 20 μg/mL laminin (Sigma, Cat. No. L2020 or equivalent). Add 5 mL laminin solution to a 75 cm2 flask and incubate overnight at room temperature. Remove laminin solution and allow flask to air dry uncapped and standing upright in a biological cabinet before introducing cells.
  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 to 10 minutes.
  4. Resuspend cell pellet with the recommended complete medium (see the specific batch information for the culture recommended dilution ratio) and dispense into the appropriate size poly-D-lysine BioCoatÒ Cellware culture flask pre-coated with laminin.  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

Volumes used in this protocol are for 75 cm2. flasks; proportionally reduce or increase amount of solutions for culture vessels of other sizes.


Note: These cells are cultured on flasks coated with 0.1 mg/mL Poly-L-Lysine (Sigma catalog # P4707 or equivalent), and then additionally coated with 20 μg/mL of Laminin (Sigma catalog # L2020 or equivalent). Add 5 mL Poly-L-Lysine solution to a 75 cm2 flask and gently rock the flask for 1 to 3 hours at room temperature. Remove the Poly-L-Lysine solution and allow flask to air dry uncapped in a biological safety cabinet. When dry, add 5 mL Laminin solution to the flask and gently rock the flask for 1 to 3 hours at room temperature. Remove the Laminin solution and allow flask to air dry uncapped in a biological safety cabinet before introducing cells. It is recommended to coat the vessel immediately prior to use. 
  1. Remove and discard culture medium.
  2. Briefly rinse the cell layer with Ca++/Mg++ free Dulbecco's 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).
    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 150 to 400 x g for 8 to 12 minutes. Discard supernatant.
  6. Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new poly-L-lysine and laminin coated culture vessels. An inoculum of 2 X 104 to 5 X 104 viable cells/cm2 is recommended.
  7. Incubate cultures at 37°C. subculture when cultures reach a cell concentration between 1 X 105 and 2 X 105 cells/cm2

Subcultivation ratio: A subcultivation ratio of 1:3 to 1:8 is recommended.
Medium renewal: Every 2 to 3 days.
Reagents for cryopreservation
Complete Culture Medium + 5% DMSO (ATCC 4-X)

Quality control specifications

Mycoplasma contamination
Not detected
Population doubling time
Approximately 50 hrs

History

Depositors
J Gordon and J Ippolito
Year of origin
2002

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

Frequently Asked Questions

References

Curated Citations

Ippolito JE, et al. An integrated functional genomics and metabolomics approach for defining poor prognosis in human neuroendocrine cancers. Proc. Natl. Acad. Sci. 102(28):9901-9906, 2005. PubMed: 15998737

Hu Y, et al. RNA interference of achaete-scute homolog 1 in mouse prostate neuroendocrine cells reveals its gene targets and DNA binding sites. Proc. Natl. Acad. Sci. 101(15):5559-5564, 2004. PubMed: 15060276

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