hTERT Human Liver Cancer-Associated Fibroblasts (CAF)

Catalog Number: HT16
Product Format: Frozen Vial
Cell Number: 1,000,000 cells/vial
Suggested Medium: CAF Growth Medium

General Information

Puromycin-resistant hTERT cells were selected from HLCCAF following lentiviral infection expressing hTERT.
These Human Liver Cancer-Associated Fibroblasts (CAFs) were isolated from human liver tumor tissue and cultured in T75 flasks coated with AlphaBioCoat. After 3–7 days in CAF Growth Medium, cells were detached at passage 1, cryopreserved, and prepared for distribution. Each vial contains at least 1,000,000 viable cells.

Product Testing

• Negative for: von Willebrand Factor/Factor VIII, cytokeratin 18, and alpha-smooth muscle actin

• Free from: bacteria, yeast, fungi, and mycoplasma

• CAF Marker Expression: FAP, PDGFR, Vimentin, PDPN, CD70

• Expansion: 3–5 passages at a split ratio of 1:2 or 1:3

Laboratory Applications

• Cell-cell interaction and adhesion assays

• PCR, Western blot, and immunoprecipitation

• Immunofluorescence and flow cytometry 

• Generation of fibroblast derivatives for oncology research

Shipping

Frozen vials on dry ice

Post-Thaw Culture Instructions

1. Thawing and Decontamination

   • Thaw vial in a 37°C water bath (2 minutes, gentle agitation).

   • Keep the O-ring and cap out of the water.

   • Spray or dip in 70% ethanol.

   • Perform all following steps under aseptic conditions.

2. Cell Recovery

   • Centrifuge cells at 125 × g for 5–10 minutes.

   • Discard supernatant and resuspend in fresh growth medium.

   • Prepare flask by coating with 6–8 mL AlphaBioCoat for 15 minutes, rinse with 8 mL 1× PBS, and discard.

   • Transfer cells into prepared T-flask.

3. Incubation

   • Incubate at 37°C in a 5% CO₂ humidified incubator.

   • To prevent alkalinity, pre-warm medium in the incubator for 15 minutes (target pH: 7.0–7.6).

Subculturing Protocol

Volumes below are for T-75 flasks. Adjust accordingly for other formats.

1. Remove spent medium.

2. Rinse cells with 1× PBS to remove residual serum.

3. Add 2.0–3.0 mL Cell Detachment Solution. Incubate 5–15 minutes at 37°C.

   • Do not tap or shake the flask.

   • For difficult detachment, extend incubation or rewarm.

4. Add 6.0–8.0 mL AlphaBioCoat to a new flask for 15 minutes, rinse with 8 mL PBS, and discard.

5. Add 6.0–8.0 mL fresh complete medium and aspirate cells gently.

6. Plate into new culture vessels.

• Subcultivation Ratio: 1:2 to 1:3

• Medium Renewal: Every 3–4 days

Cryopreservation

• Use complete growth medium supplemented with 5% (v/v) DMSO

• Catalog #: CGM5

Product Usage

Cells are offered for Research Use Only. Not for Clinical Use.