Adenovirus purification kit

The mouse hepcidin-expressing plasmid was constructed by inserting the full-length coding region of hepcidin (ID:84506) into pCDNA 3.0 vector, and then cut from pCDNA 3.0 ligated into Ad-track vector (Ad-hepcidin) and sequenced to confirm.
For gene transfer, adenovirus generation, amplification and titration were performed. Viral particles were purified using the Adenovirus Purification Kit (Cell Biolabs, San Diego, CA, USA). Min6 cells were infected with adenovirus at a multiplicity of infection of 50 at 37°C and, 2 h after infection, the cells were cultured in fresh medium for another 18 h before treating with glucose (Sigma-Aldrich) treatment. Use Ad-Gfp virus as control.

The conditions used for the transduction of recombinant adenoviruses were optimized by using adenovirus encoding GFP. All reagents and kits, including transduction reagents, an adenovirus purification kit, and an adenovirus titration kit, were purchased from Cell Biolabs, Inc. After purification, the titration of each recombinant adenovirus was determined by an ELISA titrating kit. HUVECs were seeded into 6-well plates for 24 h until they reached 80% confluence. According to the manufacturers’ protocol, adenovirus was transduced into cells by using ViraDuctin (Cell Biolabs, Inc.). HUVECs were infected with adenoviral vectors with a multiplicity of infection (MOI) of 100 plaque-forming units per cell in the presence of ViraDuctin. After incubation with viral particles for 48 h, the cells were assessed for the expression of the transduced genes.

The conditions used for the transduction of recombinant adenoviruses were optimized by using adenovirus encoding GFP. All reagents and kits, including transduction reagents, an adenovirus purification kit, and an adenovirus titration kit, were purchased from Cell Biolabs, Inc. After purification, the titration of each recombinant adenovirus was determined by an ELISA titrating kit. HUVECs were seeded into 6-well plates for 24 h until they reached 80% confluence. According to the manufacturers’ protocol, adenovirus was transduced into cells by using ViraDuctin (Cell Biolabs, Inc.). HUVECs were infected with adenoviral vectors with a multiplicity of infection (MOI) of 100 plaque-forming units per cell in the presence of ViraDuctin. After incubation with viral particles for 48 h, the cells were assessed for the expression of the transduced genes.