Adenoviruses encoding‐galactosidase (Adβ‐gal), full‐length human APE1/Ref‐1 (Ad‐APE1/Ref‐1), or PPTLS‐APE1/Ref‐1 (Ad‐PPTLS‐APE1/Ref‐1) with secretory signal sequences were prepared according to previously described methods,
16 (link),
17 (link),
18 (link) and their treatment effects were compared. Human embryonic kidney‐293 T cells (HEK293T) with the large T‐antigen of Simian virus 40 (ATCC, Manassas, VA, USA) were used to amplify and purify the adenoviruses. HEK293T cells were seeded in plates at 2.5 × 105 cells/mL and maintained at 37°C in a humidified incubator with 5% CO2. HEK293T cells were infected with 500 multiplicity of infection (particle‐forming units per cell) of the adenovirus containing Adβ‐gal, Ad‐APE1/Ref‐1, or Ad‐PPTLS‐APE1/Ref‐1 and incubated for 48 h. The adenovirus was purified using the Adeno‐X maxi Purification Kit (Takara Bio, San Jose, CA, USA). The cell culture‐amplified adenovirus was quantified using an Adeno‐X rapid titre kit (Takara Bio). APE1/Ref‐1 sandwich enzyme‐linked immunosorbent assay (ELISA; MediRedox, Daejeon, Korea) was used to quantify secretory APE1/Ref‐1 in the culture supernatant or whole‐cell lysates, as previously described.
19 (link)
16 (link),
17 (link),
18 (link) and their treatment effects were compared. Human embryonic kidney‐293 T cells (HEK293T) with the large T‐antigen of Simian virus 40 (ATCC, Manassas, VA, USA) were used to amplify and purify the adenoviruses. HEK293T cells were seeded in plates at 2.5 × 105 cells/mL and maintained at 37°C in a humidified incubator with 5% CO2. HEK293T cells were infected with 500 multiplicity of infection (particle‐forming units per cell) of the adenovirus containing Adβ‐gal, Ad‐APE1/Ref‐1, or Ad‐PPTLS‐APE1/Ref‐1 and incubated for 48 h. The adenovirus was purified using the Adeno‐X maxi Purification Kit (Takara Bio, San Jose, CA, USA). The cell culture‐amplified adenovirus was quantified using an Adeno‐X rapid titre kit (Takara Bio). APE1/Ref‐1 sandwich enzyme‐linked immunosorbent assay (ELISA; MediRedox, Daejeon, Korea) was used to quantify secretory APE1/Ref‐1 in the culture supernatant or whole‐cell lysates, as previously described.
19 (link)