Immortalized Mouse Primary Colonic Fibroblasts

Description

Maintaining a consistent mouse colonic fibroblast model across extended experiments can be limited by the finite expansion capacity of primary cells. Immortalized Mouse Primary Colonic Fibroblasts provide a C57BL/6 mouse colon-derived fibroblast model that can be expanded for more than 10 passages under the specified culture conditions.

These immortalized mouse colonic fibroblasts are derived from primary cells isolated from the colonic tissues of pathogen-free laboratory C57BL/6 mice. The cells are cultured in gelatin-precoated tissue culture flasks with Cell Biologics' Complete Growth Medium and harvested at passages 2–3 for cryopreservation.

Each vial contains 0.5 × 10⁶ cells per mL and is delivered frozen. The cells exhibit spindle morphology and are characterized by immunofluorescence staining with anti-FSP1/S100A4 antibodies. Documented microbial quality control results are negative for bacteria, yeast, fungi, and mycoplasma.

Cell Biologics' immortalized cells are derived from primary cells with an extended lifespan through overexpression of SV40 large T antigen or human telomerase reverse transcriptase (hTERT). The supplied product documentation does not identify which of these immortalization methods applies specifically to Catalog No. C57-6231IM.

Product fit: C57BL/6 mouse colon-derived fibroblasts with spindle morphology, FSP1/S100A4 characterization, cryopreserved delivery, and documented expansion for more than 10 passages.

Specifications

Features

• Derived from colonic tissues of pathogen-free laboratory C57BL/6 mice
• Primary cell-derived immortalized fibroblasts with an extended culture lifespan
• Documented expansion for more than 10 passages under the specified conditions
• Harvested at passages 2–3 and supplied as a cryopreserved vial
• Contains 0.5 × 10⁶ cells per mL
• Characterized by spindle morphology and anti-FSP1/S100A4 immunofluorescence staining
• Tested negative for bacteria, yeast, fungi, and mycoplasma
• Recommended for subculture at a split ratio of 1:2
• Supported by a complete fibroblast medium and gelatin-based coating workflow
• Shipped frozen on dry ice for transfer to liquid nitrogen storage upon arrival

Applications

Immortalized Mouse Primary Colonic Fibroblasts are intended for in vitro research requiring a C57BL/6 mouse colon-derived fibroblast model with an extended culture lifespan. Their documented spindle morphology, FSP1/S100A4 immunofluorescence characterization, and expansion for more than 10 passages help researchers evaluate whether the cells match the identity and culture-duration requirements of a planned study.

Research uses supported by the documented product characteristics include:

• In vitro culture of C57BL/6 mouse colonic fibroblasts
• Studies requiring a mouse colon-derived fibroblast culture model
• Observation of fibroblast spindle morphology
• FSP1/S100A4-associated fibroblast characterization
• Cell growth, passage, and expansion studies
• Experiments requiring continued fibroblast expansion for more than 10 passages
• Comparative studies requiring cryopreserved mouse colonic fibroblast cells
• General cell-based assays performed under the specified fibroblast culture conditions

Application selection should be based on the documented cell origin, characterization, and culture properties. The supplied documentation does not claim product-specific validation for a particular disease model, therapeutic application, or diagnostic procedure.

Cell Culture Workflow

Culture Preparation

• Perform all cell culture procedures in a biosafety cabinet using aseptic technique.
• Use culture media, supplements, and reagents that have been sterile filtered through a 0.2 μm filter.
• Prewarm only the amount of culture medium required for immediate use.
• Repeated warming of complete culture medium is not recommended.
• Use M2267 Complete Fibroblast Medium w/ Kit as the suggested medium.

Gelatin Coating

• Coat sterile culture flasks or dishes with CB6950 Gelatin-Based Coating Solution.
• Allow the coating solution to contact the culture surface for 2 minutes.
• Aspirate the excess coating solution before seeding cells.

Recovery of Cryopreserved Cells

1. Thaw the cryovial rapidly in a 37°C water bath for less than 1 minute or until only a small amount of ice remains.
2. Remove the vial and disinfect its exterior with 70% ethanol.
3. Transfer the cells to a sterile 15 mL conical tube containing 5 mL of prewarmed culture medium.
4. Rinse the vial with an additional 0.5–1 mL of medium to recover remaining cells.
5. Centrifuge at 200 × g for 5 minutes to remove residual DMSO.
6. Carefully aspirate the supernatant and resuspend the cell pellet in 5 mL of prewarmed growth medium.
7. Seed the resuspended cells into a gelatin-precoated T25 flask.
8. Place the flask in a humidified 37°C incubator with 5% CO₂.
9. Leave the culture undisturbed for 12–16 hours.
10. Monitor cell confluence and morphology daily under a microscope.

Subculture

• Subculture the cells at approximately 90% confluence.
• Remove the culture medium and wash the adherent cells with calcium- and magnesium-free 1X PBS.
• Incubate the cells with prewarmed 0.25% Trypsin-EDTA Solution (Catalog No. CB6914) for 3–5 minutes.
• Once the cells detach, add complete culture medium containing serum to neutralize the trypsin.
• Collect the cell suspension and centrifuge at 200 × g for 5 minutes.
• Resuspend the cell pellet in fresh medium and seed into newly gelatin-coated culture vessels.
• Use the documented split ratio of 1:2.

The linked culture documents contain general primary-cell handling instructions. Consult the supplied protocols and technical support before beginning culture, particularly where protocol versions provide different operating volumes or medium-change intervals.

Quality Control

Storage and Handling

• Cryopreserved cells are shipped with dry ice overnight.
• Upon arrival, immediately transfer the frozen vial from the dry-ice shipping container to a liquid nitrogen storage system.
• Do not use a −20°C or −80°C freezer for routine long-term storage.
• Repeated freezing and thawing of the cells are not recommended.
• Live-cell shipment is available on request.
• Handle all animal cells with caution as potential biohazards because no testing procedure can guarantee the complete absence of infectious agents.
• Use appropriate laboratory practices and personal protective equipment when handling the product.

Documents

C57-6231IM Product Datasheet

Primary Cell Culture Protocol

Primary Cell Culture Protocol (Supplementary)

Cell Culture Products Safety Data Sheet

Frequently Asked Questions

What are Immortalized Mouse Primary Colonic Fibroblasts?

They are primary cell-derived fibroblasts isolated from the colonic tissues of pathogen-free laboratory C57BL/6 mice and immortalized to provide an extended culture lifespan. The cells are supplied cryopreserved for research use.

Are C57-6231IM cells primary or immortalized?

C57-6231IM cells are immortalized cells derived from primary mouse colonic fibroblasts. The term “primary” describes the origin of the cells, while “immortalized” describes their extended culture lifespan.

What mouse strain and tissue are these fibroblasts derived from?

The cells are derived from the colonic tissues of pathogen-free laboratory C57BL/6 mice.

How many cells are supplied?

The product datasheet states that each cryopreserved vial contains 0.5 × 10⁶ cells per mL. The documentation does not provide a separate post-thaw viability value.

At what passage are the cells cryopreserved?

The cells are harvested from culture flasks at passages 2–3 and cryopreserved in vials.

How are the mouse colonic fibroblasts characterized?

The cells are characterized by their spindle morphology and by immunofluorescence staining with anti-FSP1/S100A4 antibodies.

What quality control testing is documented?

The product datasheet reports negative results for bacteria, yeast, fungi, and mycoplasma.

How many passages can C57-6231IM cells be expanded?

Under the cell culture conditions specified by Cell Biologics, the immortalized mouse colonic fibroblasts can be expanded for more than 10 passages at a recommended split ratio of 1:2.

What medium is recommended for C57-6231IM cells?

The suggested medium is M2267 Complete Fibroblast Medium w/ Kit – 500 mL.

Do the culture vessels require coating?

Yes. The supplied documentation specifies gelatin-precoated culture vessels. CB6950 Gelatin-Based Coating Solution is used to coat the culture surface for 2 minutes before excess solution is aspirated.

What culture conditions are recommended?

The supplied culture protocol specifies a humidified incubator at 37°C with 5% CO₂. The documented split ratio is 1:2.

Which immortalization method is used for C57-6231IM?

The product documentation states that Cell Biologics' immortalized cells are generated through overexpression of SV40 large T antigen or human telomerase reverse transcriptase (hTERT). It does not identify which method applies specifically to C57-6231IM. Contact technical support for product-specific confirmation.

How should the cells be handled after delivery?

Cryopreserved cells are shipped on dry ice. Upon arrival, immediately transfer the frozen vial to a liquid nitrogen storage system until the cells are ready for use.

Can the cells be stored in a −80°C freezer?

No. The product documentation instructs users to transfer the cells to liquid nitrogen immediately upon arrival and states that a −20°C or −80°C freezer should not be used for routine storage.

Is repeated freezing and thawing recommended?

No. Repeated freezing and thawing of the cells are not recommended.

Are these cells intended for clinical or diagnostic use?

No. Immortalized Mouse Primary Colonic Fibroblasts are for research use only and are not authorized for human use, in vitro diagnostic procedures, or therapeutic procedures.