HEK293

HEK 293 cells (also written HEK293, HEK-293, or 293 cells) are an immortalized human cell line created in 1977 by transforming cultured human embryonic kidney cells with sheared DNA from human adenovirus type 5 (Ad5). The line, deposited by Frank L. Graham of McMaster University, is held as ATCC CRL-1573 and is one of the most widely used human cell lines in biomedical research and biomanufacturing.^{1 • 2}

Creation and transformation

The line was established by exposing human embryonic kidney cells to sheared fragments of adenovirus type 5 DNA. The transformed cells, designated 293, showed characteristics of transformation including production of a virus-specific tumor antigen, and hybridization experiments showed transcription of adenoviral sequences from the left-hand end of the viral genome.¹ The Ad5 insert was later cloned and sequenced, establishing that a colinear segment from nucleotides 1 to 4344 is integrated into chromosome 19 at 19q13.2, within the pregnancy-specific beta-1-glycoprotein 4 (PSG4) gene.^{2 • 7} The integrated left-end sequences drive constitutive expression of the adenoviral E1 genes, which is why the cells support replication of E1-deleted adenoviral vectors.⁷

The two major cell banks describe the viral content differently. ATCC states that an earlier report suggesting DNA from both ends of the Ad5 genome was wrong and that only left-end sequences are present.² The German Collection of Microorganisms and Cell Cultures (DSMZ) describes the cells as containing 4 to 5 copies of the left end of the virus (12% of the viral genome, including E1a and E1b) and one copy of the right end (10% of the genome, including the E4 gene).⁸

Genome and karyotype

293 cells carry an aberrant, hypotriploid karyotype with a modal chromosome number of 64. ATCC's characterization records marker chromosomes including der(1)t(1;15)(q42;q13) and der(19)t(3;19)(q12;q13), three copies of the X chromosome in most cells, and four copies of chromosomes 17 and 22. The line is tumorigenic, forming tumors in nude mice.² Continuously cultured immortalized lines of this kind show a high degree of genomic instability with frequent chromosome rearrangements, so different HEK293 lineages propagated in different laboratories differ at the genomic and transcriptomic level from the parental line.⁹ Resequencing of six 293 cell lines has identified single-nucleotide polymorphisms common to the lineage, intended to support future cell line authentication assays.³

Cell of origin

Although the line is named for embryonic kidney, the tissue the cells most resemble has been questioned. A genome and transcriptome analysis intersected the 293 transcriptome with cell-type signatures from kidney, central nervous tissue, adrenal tissue, and pituitary tissue, and concluded that an adrenal lineage is the most likely among those tested. The authors proposed that the cells arose in the embryonic adrenal precursor structure, which lies adjacent to the kidney during development; the adrenal medulla is of neural crest ectodermal origin, which could explain the expression of some neuron-specific genes in the line.³ The aberrant karyotype, genomic heterogeneity, and constitutive adenoviral E1 expression severely deregulate the transcriptome, making it difficult to identify an authentic cell type of origin.⁷

Uses in research and manufacturing

293 cells and their derivatives are used as a heterologous expression system for recombinant proteins and adenovirus production, valued for human-like post-translational modification of proteins, high transfection efficiency, fast growth, and growth in serum-free suspension culture.^{5 • 6} Applications span biopharmaceutical manufacturing, drug screening, structural biology, and electrophysiology; the line and its derivatives have been among the most frequently used cells in cell biology research after HeLa, and in biotechnology after CHO.^{5 • 7}

Vector manufacturing. Transfection of plasmid DNA into HEK293 cells remains the most commonly used protocol for clinical-grade manufacturing of recombinant adeno-associated virus (AAV) vectors, using adherent CellSTACK cultures or suspension cultures in shaker flasks and WAVE bioreactors with two- or three-plasmid transfection.¹⁰ HEK293-derived suspension lines such as HEK 293SF-3F6, HEK 293FT, and SJ293TS have been developed for lentiviral vector production and have achieved titers of around 10^8 transducing units per ml.¹¹ Virus-like particles for vaccines, including candidates against rabies, influenza A H1N1, and hepatitis B, have also been produced in the line.⁴

Approved therapeutics. Several approved biologics are made in HEK293 derivatives: recombinant FVIIIFc and FIXFc (ALPROLIX, 2014, and ELOCTATE) in HEK293-H, human-cl rhFVIII (NUWIQ) in HEK293F, and the GLP-1 Fc fusion protein dulaglutide (TRULICITY) in HEK293 EBNA; the withdrawn drotrecogin alfa (XIGRIS) was produced in the parental line.⁴ Gene therapy products using HEK293-made vectors include the lentiviral vectors Kymriah and Zynteglo, the AAV vectors Glybera, Luxturna, and Zolgensma, and retroviral vectors such as Rexin-G and Strimvelis.⁴

Comparison with CHO cells

CHO cells dominate industrial protein production, with approximately 80% of approved recombinant therapeutic proteins made in them, and stable CHO lines remain the standard for manufacturing biopharmaceuticals.^{6 • 12} HEK293's advantages are its human origin, high transfection efficiency, fast growth, and suitability for serum-free suspension culture, which allow high-quality, biologically active recombinant proteins.⁶ In a side-by-side expression test of 24 difficult-to-express human secreted proteins, one third showed improved secretion when the host was switched from CHO to HEK293; 8 of 24 genes expressed better in HEK293 in both semi-stable and transient setups, while only PLG expressed better in CHO, and four proteins were detected only in HEK293 supernatants.¹³

The glycosylation profiles of the two hosts differ. Glycoproteins expressed in HEK293 cells generally show more complex glycosylation profiles than CHO-derived proteins, but HEK293 products carry high levels of terminal N-acetylgalactosamine (GalNAc) modification and low levels of sialylation, which complicates their use for therapeutic proteins.^{14 • 12} Because obtaining homogeneous glycosylation patterns is practically impossible without genetic manipulation or chemical treatment, most glycosylation engineering in HEK293 targets glycan-processing enzymes. Two engineered lines are in use: HEK293SG, in which the N-acetylglucosaminyltransferase I (GnTI) gene is knocked out so the cells lack complex N-glycans, and its derivative HEK293SGGD (293SGlycoDelete), which expresses a heterologous Golgi endoglycosidase to secrete de-glycosylated glycoproteins. Small-molecule inhibitors such as kifunensine, which targets alpha-mannosidase I, and swainsonine, which blocks alpha-mannosidase II, produce chemically uniform glycans.⁴

Derivatives

The best-known derivative is HEK 293T, generated by introducing a temperature-sensitive allele of the SV40 large T antigen into the 293 genome. This enables replication of plasmids containing the SV40 origin of replication, enhancing gene expression and yielding higher titers than the original line.¹¹ Other derivatives include suspension-adapted lines such as HEK293F and HEK293 EBNA used for production, the glycoengineered HEK293SG and HEK293SGGD lines, and 293T/17, a high-transfection 293T subclone distributed by ATCC.^{4 • 2} Cell line engineering targeting genes involved in proliferation, apoptosis, metabolism, glycosylation, secretion, and protein folding, together with media and vector optimization, has further improved growth and production performance.¹⁵

References

1. Graham FL, Smiley J, Russell WC, Nairn R (1977). Characteristics of a Human Cell Line Transformed by DNA from Human Adenovirus Type 5. Journal of General Virology 36:59-72.
2. [293 [HEK-293] CRL-1573 product page, ATCC](https://www.atcc.org/products/cells_and_microorganisms/by_tissue/kidney/crl-1573)
3. Lin Y-C et al. Genome dynamics of the human embryonic kidney 293 lineage in response to cell biology manipulations. PMC.
4. Affecting HEK293 Cell Growth and Production Performance by Modifying the Expression of Specific Genes. Cells 10:1667 (2021), PMC.
5. Human Embryonic Kidney 293 Cells: A Vehicle for Biopharmaceutical Manufacturing, Structural Biology, and Electrophysiology. PubMed.
6. Zhang L et al. Current strategies for the development of high-yield HEK293 cell lines. Biochemical Engineering Journal 205 (2024).
7. HEK293 in cell biology and cancer research: phenotype, karyotype, tumorigenicity, and stress-induced genome-phenotype evolution. ScienceDirect.
8. 293 (ACC 305), Leibniz Institute DSMZ
9. Evolution from adherent to suspension: systems biology of HEK293 cell line development. PMC.
10. Manufacturing of recombinant adeno-associated viral vectors for clinical trials. PMC.
11. Ripoll Fiol, EngD thesis, University College London (HEK 293T and lentiviral vector producer lines)
12. State-of-the-Art Approaches to Heterologous Expression of Bispecific Antibodies Targeting Solid Tumors. Biochemistry (Moscow), Springer.
13. Harnessing secretory pathway differences between HEK293 and CHO to rescue production of difficult to express proteins. PMC.
14. Impact of host cell line choice on glycan profile. Taylor & Francis.
15. Affecting HEK293 Cell Growth and Production Performance by Modifying the Expression of Specific Genes. Cells 10:1667 (2021), MDPI.

Topic: Encyclopedia › Life and health › Biological foundations › Development and comparative physiology › Cellular, regenerative, and comparative physiology › Embryonic and adult stem cells › Established stem cell lines

Initially written Sep 17, 2026 · Reviewed: Oct 21, 2026 · Edited: Oct 15, 2026; Oct 21, 2026 · Last review: Oct 21, 2026

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