Catalog Number GF9Product Name GFP Expressing human liver sinusoidal endothelial cells(GFP- HLSECs)Storage Liquid NitrogenProduct Format Frozen vialCells Number >90% confluent in Frozen Vial*Caution: The handling of human derived products has the potential to be biologically hazardous. All Cell strains tested negative for HIV, HBV,and HCV DNA in diagnostic tests. Proper precautions must be taken to avoid exposure. Always wear proper protective equipment (Gloves,safety glasses, etc.) when handling these materials. We recommend following the universal procedures for handling products of human originas the minimum precaution against contamination.GENERAL INFORMATIONGFP expressing human liver sinusoidal endothelial cells were initiated by elutriation from dissociatednormal human liver tissue and transfected with GFP- lentiviral particles at passage 1. Puromycinresistant GFP-HLSECs were selected and shipped in proliferating culture or frozen vial with a confluenceof > 90% (cells are providing at passage 4-5). ENDO-Growth medium (EGM-2102) containing 5% serumand growth supplement is recommended for culture. Cells have an average population doubling level>16 when cultured. When you receive the cells, leave the flask in 37°C C0₂ Incubators for 1 hour. Then,replace the transport medium with fresh ENDO-Growth medium (EGM-2102). Let the cells grow for 24hours before subculture.CELL CHARACTERIZATIONCytoplasmic VWF/ factor VIII >95% positive by immunofluorescenceCytoplasmic uptake of Di-I-Ac-LDL >95% positive by immunofluorescenceCytoplasmic PECAM1 >95% positive by immunofluorescenceGFP Expressing human liver sinusoidalendothelial cells are negative forHIV-1, HBV, HCV, and mycoplasmaPRODUCT USE AND SHIPPING STATUSProduct Use GFP Expressing human liver sinusoidalendothelial cells are for research use onlyShipping Status Frozen via
Frozen1) Coating T25 flasks. Add 2 ml AlphaBioCoat (AC001) into a T25 flask and ensure entire interiorsurface is coated with the solution. After 30 minutes, dispose of AlphaBioCoat (AC001) byaspiration. Gently rinse and aspirate flask with Phosphate Buffer Solution (1XPBS-001). The flaskis now ready for use (no need for overnight incubation when coated with AC001).2) If you are using the coated flask the same day, add about 4 ml of Endo-Growth media(EGM2102) to the coated flask. If the media changes color from pink to yellow, aspirate and discardthe media. Add 4ml of fresh media to the coated flask.3) Thaw the cells in a 37°C water bath. Once you see a small amount of ice left in the vial, spray thevial with 70% Ethanol and wipe it down.4) Transfer the vial into your Biosafety cabinet.5) Using a 2 or 5ml pipet, pipet the cells out of the vial.6) Transfer your cell suspension in to your coated plate that have the 4 ml media in it.7) You should have a total working volume of 5ml of cell suspension in the flask; close the cap.Make sure cells are evenly distributed in the flask by moving the flask left and right five times.Move it up and down for and additional five times.8) Place flask in a 37°C incubator with 5% C02. If flask is not vented, please loosen cap.9) Change media after 48 hours.