Human Colorectal Tumor CAFs

Product Information

Catalog Number: HC-CA01
Applications: Cell Assays
Type: Human Primary CAFs
Organism: Homo sapiens (human)
Format: Frozen
Storage: Keep frozen in liquid nitrogen until plating
Shipping: Frozen (Dry Ice / Liquid Nitrogen)

Note: Cultures should be stored in the vapor phase of liquid nitrogen, not submerged.

Handling Information

Unpacking & Storage Instructions

1. Check all containers for leakage or breakage.

2. Remove the frozen cells from dry ice and immediately store at below -130°C, preferably in liquid nitrogen vapor phase.

Culture Medium & Conditions

• Complete Medium: Obtain the base medium and additives from our company.

• Temperature: 37°C

• Atmosphere: 5% CO₂ in air recommended

Handling Procedure

Important: To ensure maximum cell viability, thaw and culture the cells immediately upon receipt.

If storage is necessary, do not store at -70°C — use the vapor phase of liquid nitrogen instead.

1. Thaw the vial quickly in a 37°C water bath (approx. 2 minutes). Keep the cap and O-ring out of water.

2. Once thawed, decontaminate the vial by spraying or dipping in 70% ethanol.

   • Perform the remaining steps under aseptic conditions.

3. Transfer contents to a centrifuge tube with 9.0 mL of complete low-serum culture medium.

   • Centrifuge at ~125 x g for 5–10 minutes.

   • Discard the supernatant.

4. Resuspend the cell pellet in complete medium at the recommended dilution.

   • Use a 75 cm² tissue culture flask.

   • Before adding cells, pre-incubate the medium in the flask for at least 15 minutes to allow pH to normalize (7.0–7.6).

5. Incubate at 37°C with 5% CO₂ atmosphere.

Subculturing Procedure

Quantities below are for a 75 cm² flask. Adjust proportionally for other flask sizes.

1. Remove and discard old culture medium.

2. Rinse the cell layer briefly with 1x PBS, then discard the PBS.

3. Add 2.0 to 3.0 mL of cell detachment solution.

   • Observe under a microscope.

   • Cells should detach within 5–15 minutes.

   • Do not agitate the flask to avoid clumping.

   • Incubate at 37°C if cells are slow to detach.

4. Add 6.0 to 8.0 mL of complete growth medium and gently pipette to aspirate cells.

5. Transfer to centrifuge tube. Spin at ~125 x g for 5–10 minutes.

   • Discard supernatant.

   • Resuspend in fresh serum-free medium.

   • Plate into new culture vessels.

6. Incubate at 37°C.

• Subcultivation Ratio: 1:2 to 1:6

• Medium Renewal: Every 2 to 3 days