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HEK 293T, a highly transfectable derivative of the parental HEK 293 cell, stands out as a versatile and powerful tool in the field of biotechnology for the production of recombinant proteins and various types of vaccines.
HEK-293T cells were generated by transfecting embryonic kidney 293 cells with a plasmid encoding the SV40 large T antigen. The original HEK293 cell line was developed from the epithelial cells of human embryonic kidney tissue, with its transformation occurring in what was notably the 293rd experiment conducted by the researchers.
In the realm of vaccine development, the 293T embryonic kidney cells are pivotal for viral vector production, including adenovirus vectors. HEK293T cells, under specific culture conditions, are transfected with vectors carrying adenoviral and retroviral elements, including the SV40 origin of replication, leading to the production of virus-like particles (VLPs).
The VLPs, devoid of viral genetic material, are key in forming the basis of subunit and VLP-based vaccines. The recombinant protein production in 293T cells is facilitated by various transfection methods, with an emphasis on the generation of AP fusion proteins and other protein types that form the antigenic component of vaccines.
The 293T cell line's genome engineering capabilities allow for the customization of expression constructs, further boosting the production of viral vectors. This, coupled with the ability to produce proteins in suspension culture or adherent conditions, makes the 293T cell line a full-stack solution for modern vaccine development.
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- Receptors expressed: Vitronectin
- Protein expression: CEA negative, p53 positive
- Tumorigenic: In nude mice
- cultureMedium: EMEM (MEM Eagle), w: 2 mM L-Glutamine, w: 2.2 g/L NaHCO3, w: EBSS (Cytion article number 820100a)
- supplements: Supplement the medium with 10% FBS and 1% NEAA
- dissociationReagent: Accutase
- doublingTime: 30 hours
- subculturing: Remove the old medium from the adherent cells and wash them with PBS that lacks calcium and magnesium. For T25 flasks, use 3-5 ml of PBS, and for T75 flasks, use 5-10 ml. Then, cover the cells completely with Accutase, using 1-2 ml for T25 flasks and 2.5 ml for T75 flasks. Let the cells incubate at room temperature for 8-10 minutes to detach them. After incubation, gently mix the cells with 10 ml of medium to resuspend them, then centrifuge at 300xg for 3 minutes. Discard the supernatant, resuspend the cells in fresh medium, and transfer them into new flasks that already contain fresh medium.
- seedingDensity: 1 x 104 cells/cm2 will yield in a confluent layer in about 4 days
- fluidRenewal: 2 times per week
- postThawRecovery: After thawing, plate the cells at 5 x 104 cells/cm2 and allow the cells to recover from the freezing process and to adhere for at least 24 hours.
- freezeMedium: As a cryopreservation medium, use complete growth medium (including FBS) + 10% DMSO for adequate post-thaw viability, or CM-1 (Cytion catalog number 800100), which includes optimized osmoprotectants and metabolic stabilizers to enhance recovery and reduce cryo-induced stress.
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