|Name of Cells:
||GFP Human Lung Fibroblasts (HLFCs-Ad)
HLFCs-Ad are isolated from healthy human lung tissue (adult) samples and shipped in frozen vials (the cells are provided @ passage 1). DMEM contains 10% Fetal Bovine Serum (Full medium) is recommended for cell culture and these cells have an average minimum population doubling levels > at least 14 when cultured following the detailed protocol described below). HLFCs-Ad are tested negative for HIV-1, HBV, HCV, and mycoplasma.
Product Use: HLFCs-Ad are for research use only.
Shipping: Frozen Vial.
Handling of Arriving Cells
When you receive the cells in a frozen vial, you can transfer the vial of cells into a -80ºC freezer for short term storage or a liquid nitrogen tank for long term storage. Thaw the cells in a 37°C water bath, and then transfer the cells into a T25 flask pre-coated with Universal Coating Solution (UBP-01) as described in detail in Subculture Protocol.
- Pre-coating of T25 flasks: Add 2ml of Universal Coating Solution (UBP-01) into one T25 flask and make sure whole surface of the flask is covered with the coating solution. Five minutes later, dispose excessive Universal Coating Solution by aspiration and the flask is ready to be used (no need for overnight incubation when using Universal Coating Solution).
- Rinse the cells in T25 flask with 5ml HBSS (Room Temperature, RT) twice.
- Add 2ml of Universal Detachment Solution (RT) (UBP-23) into one T25 flask (make sure the whole surface of the T25 flask is covered with Universal Detachment Solution), and gently dispose the excessive Universal Detachment Solution within 20 seconds with aspiration.
- Leave the T25 flask with the cells at RT for 1 minute (the cells usually will detach from the surface within 1-2 minutes). You can monitor the cells under microscope and when most of cells become rounded up, hit the flask against the bench surface, and the cells will move on the surface of the flask when monitoring under microscope.
- Add 5ml Universal Neutralization Buffer (UBP-28) and spin the cells down with 800g for 5 minutes.
- Re-suspend the cell pellet with 10-15ml of full medium and the cell suspension is transferred directly into 2 or 3 pre-coated T25 flasks (5ml each, and the cells are sub-cultured at 1:2 to 1:3 ratios)
Change medium every 2-3 days and cells usually become confluent within 7 days (when split at a 1:3 ratio).