Immortalized Human Pancreatic Fibroblast Cells

Catalog Number: HPF001
Product Format: Frozen Vial
Cell Number: > 5 × 10⁵ cells/vial
Product Usage: Research Use Only

General Information

These puromycin-resistant hTERT-immortalized cells were derived from HPCAF after infection with lentiviruses expressing human telomerase reverse transcriptase (hTERT).
The cell line exhibits fibroblast morphology and was isolated from normal human pancreatic tissue. Cells are provided at passage 2 in frozen vials.

Shipping

Frozen vials shipped on dry ice.

To ensure optimal viability, thaw and culture immediately upon receipt. If storage is required, maintain vials in liquid nitrogen vapor phase (not at -70°C).
Storage at -70°C will result in cell death.

Thaw the vial by gentle agitation in a 37°C water bath (approx. 2 minutes).
To reduce contamination risk, keep the O-ring and cap out of the water during thawing.

Thawing and Initiation Procedure

1. Immediately after thawing, decontaminate the vial with 70% ethanol. Perform all subsequent steps under strict aseptic conditions.

2. Remove the cryoprotective agent by centrifuging the cell suspension at 125 × g for 5–10 minutes. Discard the supernatant and resuspend the pellet in fresh growth medium.

3. Add 6.0–8.0 mL of AlphaBioCoat to a T-75 flask and incubate for 15 minutes. Aspirate and rinse with 8 mL of 1× PBS, then discard the PBS. Transfer cells to the coated flask.

4. To avoid excessive alkalinity, pre-incubate the medium at 37°C in the incubator for at least 15 minutes prior to cell transfer, allowing the pH to stabilize (target range: 7.0 to 7.6).

5. Incubate the culture at 37°C in a humidified incubator with 5% CO₂ in air.

Subculturing Procedure

Instructions below are for a T-75 flask; adjust volumes proportionally for other flask sizes.

1. Remove and discard the old culture medium.

2. Briefly rinse the cell layer with 1× PBS to eliminate serum traces.

3. Add 2.0–3.0 mL of Cell Detachment Solution. Observe under an inverted microscope until the monolayer is dispersed (5–15 minutes).

   • Do not shake or tap the flask; this can cause clumping.

   • For difficult detachment, place the flask at 37°C to aid dispersal.

4. Add 6.0–8.0 mL of AlphaBioCoat to a new T-Flask and incubate for 15 minutes. Rinse with 8 mL of 1× PBS, discard, and transfer the cell suspension.

5. Add 6.0–8.0 mL of complete growth medium and gently pipette to aspirate the cells.

6. Plate appropriate aliquots into new culture vessels.

7. Incubate at 37°C.

• Subcultivation Ratio: 1:2 to 1:8

• Medium Renewal: Every 3 to 4 days

Cryopreservation Reagents

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

Product Usage

Cells are offered for Research Use Only.