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| Name of Product | GFP-Human Lung Fibroblasts (HLFCs)-Adults |
| Catalog Number | cAP-0033GFP |
| Product Format | Frozen Vial |
| Cell Number | >5x10[5]cells/vial |
| HLFCs-Ad are isolated from healthy human lung tissue (adult) samples and GFP-Human Lung Fibroblasts (GFP-HLFCs-ad) were selected by GFP-lentiviral infected human lung fibroblasts with 4ug/ml of puromycine and shipped in frozen vials (the cells are provided @ passage 3). Fibroblast Growth Medium (FGM) (cAP-44) is recommended for cell culture and these cells have an average minimum population doubling levels > at least 10 when cultured following the detailed protocol described below). HLFCs-Ad are tested negative for HIV-1, HBV, HCV, and mycoplasma. |
| Product Use | GFP-HLFCs-Ad are for research use only. |
| Shipping | Frozen Vial |
| When you receive the cells in a frozen vial, you can transfer the vial of cells into a -80C freezer for short term storage or a liquid nitrogen tank for long term storage. Thaw the cells in a 37C water bath, and then transfer the cells into a T25 flask pre-coated with Quick coating solution (cAP-01) as described in detail in Subculture Protocol. |
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| A) | Pre-coating of T25 flasks- Add 2ml of Quick Coating Solution (cAP-01) into one T25 flask and make sure whole surface of the flask is covered with the coating solution. Five minutes later, dispose excessive Quick Coating Solution by aspiration and the flask is ready to be used (no need for overnight incubation when using Quick Coating Solution). Other extracellular matrix can be used including gelatin, collagen, and fibronectin and you are advised to test the conditions for using those materials in advance. |
| B) | Rinse the cells in T25 flask with 5ml HBSS (Room Temperature, RT) twice. |
| C) | Add 2ml of Trypsin/EDTA (RT) (cAP-23) into one T25 flask (make sure the whole surface of the T25 flask is covered with Trypsin/EDTA), and gently dispose the excessive Trypsin/EDTA solution within 20 seconds with aspiration. |
| D) | 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 the 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 a microscope. |
| E) | Add 5ml Trypsin Neutralization Buffer (cAP-28) and spin the cells down with 800g for 5 minutes. |
| F) | 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 to 2 to 1 to 3 ratios). |
| G) | Change medium every 2-3 days and cells usually become confluent within 7 days (when split at a 1 to 3 ratio). |
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| Quick Coating Solution | cAP-01 240ml |
| Fibroblast Growth Medium (FGM) | cAP-44 500ml |
| HBSS w/o Ca2+ , Mg2+ | cAP-11 100ml |
| Cell Freezing Solution (FBS) | cAP-22 50ml |
| Cell Freezing Solution (Non-FBS) | cAP-22B 50ml |
| Trypsin/EDTA Solution | cAP-23 100ml |
| Trypsin Neutralization Solution | cAP-28 100ml |
| ITS (100x) | cAP-26 10ml |
| L-Glutamine-MAXIMUM (100x) | cAP-27 100ml |
| Human Plasma Fibronectin Solution | cAP-42 1mg |
