T225 CellBind cell culture flasks (10×; Corning) were seeded with approximately 5 × 106 human embryonic kidney 293 (HEK-293) T-REx cells (Invitrogen) each. Cells were cultured in Dulbecco’s modified Eagle’s medium (Gibco) supplemented with 10% heat-inactivated fetal bovine serum and 1% penicillin/streptomycin (Gibco) until 80% confluent. Cells were infected with ChAdOx1.eGFP, provided by the Coughlan Lab (University of Maryland), at a multiplicity of infection of 0.01. Cells were then monitored for cytopathic effect (CPE). When culture medium turned yellow, medium was replaced. Once CPE became evident, but the cells were not ready to be collected, yellowed medium was supplemented with sodium bicarbonate buffer (pH 7.4, Gibco) until it reddened. This was done to retain the virus released into the medium. Once CPE was observed in >80% of the cell monolayer, the cells (5 to 8 days after infection) were dissociated from the flask by knocking. The supernatant and cells were separated by centrifugation at 300g for 5 min, both were stored at −80°C until ready for purification.
Sodium bicarbonate buffer
Manufactured by Thermo Fisher Scientific
Sodium bicarbonate buffer is a chemical solution used in various laboratory applications. It serves as a buffer, maintaining a specific pH range within a solution. The core function of this product is to stabilize the pH, ensuring a consistent and controlled environment for various experiments and procedures.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
T rex hek293 cells, by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (1 mentions)
Cellbind cell culture flasks, by Corning (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
2 protocols using sodium bicarbonate buffer
1
Recombinant Adenovirus Production in HEK293 Cells
2
Propagation of ChAdOx1 Virus in HEK-293 T-Rex Cells
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Propagation of ChAdOx1 virus 10 x T225 CellBind™ cell culture flasks (Corning) were seeded with approximately 5x10 6 (link) HEK-293 T-Rex cells (Invitrogen) each. Cells were cultured in DMEM (Gibco) supplemented with 10% heat inactivated fetal bovine serum and 1% penicillin/streptomycin (Gibco) until 80% confluent. Cells were infected with ChAdOx1.eGFP, provided by the Coughlan Lab (University of Maryland), at a multiplicity of infection of 0.01. Cells were then monitored for cytopathic effect (CPE). When culture media turned yellow media was replaced. Once CPE became evident, but the cells were not ready to be collected, yellowed media was supplemented with sodium bicarbonate buffer (pH7.4, Gibco) until it reddened. This was done to retain the virus released into the media. Once CPE was observed in >80% of the cell monolayer the cells (5-8 days post infection) were dissociated from the flask by knocking. The supernatant and cells were separated by centrifugation at 300g for 5mins, both were stored at -80C until ready for purification.
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