Human TMPRSS2 knockout Caco-2 cell line

Cell culture

Biosafety level

EU: 1 US: 1

Adherent/suspension

Adherent

Gender

Male

Initial handling guidelines

Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability. <p>1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.<br>2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.<br>3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.<br>4. Incubate the culture at 37°C incubator with 5% CO₂. Check the culture one day after revival and continue to check until 70% confluent.<br>5. Once confluent passage into an appropriate flask at a density of 2x10⁴ cells/cm². Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.</p>

Subculture guidelines

• Do not allow cells to become more than 70% confluent as they will form vacuoles in the cytoplasm.
• Slow to trypsinise, however, prolonged trypsinisation is detrimental to cell viability. Trypsinise until 70% of cells have detached.
• All seeding densities should be based on cell counts gained by established methods.
• A guide seeding density of 1x10⁴ - 2x10⁴ cells/cm² is recommended.

Culture medium

EMEM + 20% FBS

Cryopreservation medium

Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.