What are HEK293 cells?

HEK293 is a cell type established by transforming human embryonic kidney cells with sheared adenovirus type 5 DNA in 1973. Due to its ease of maintenance and robustness, the fast growing cell line and its derivatives are commonly applied in biotechnology such as in the production of biopharmaceutical products and gene therapy — e. g. the expression of recombinant monoclonal antibodies –, as well as in cell biology research.

HEK293 cells — a definition

HEK293 cells are defined as immortalized human embryonic kidney cells grown in tissue culture. The cell line, often also referred to as human embryonic kidney 293, HEK-293, HEK 293 or 293 cells, is the transformation product of a miscarried female fetus of unknown origin that was exposed to sheared adenovirus 5 DNA. This is why HEK293 is not only commonly used in receptor signaling and cancer research, but also for protein production at large scale.

History of HEK293 cells

The history of HEK293 cells goes back to 1973 when Alex Van Der Eb and his team at the University of Leiden, Netherlands, cultured the cells. One of the post docs, Frank Graham, performed the transduction by adenovirus 5. The number in the cell naming can be traced back to the fact that the original cell clone resulted from his 293rd experiment. Due to the presence of Ad5 E1A/B genes, HEK293 is frequently used in the production of adenoviral vectors and adeno-associated viral vectors.

Genome, cell morphology and more — characterization of HEK293 cells

Genome

The genome of HEK293 has the characteristics of a human cell line. This also implies that it is suited for producing biotherapeutics since they allow destinctive post-translational modifications. The cells have a very complex karyotype with a chromosome number of 64, four copies of chromosome 17, three X chromosome copies, no Y chromosome-derived sequences, and an integration of 4 kilobases adenovirus type 5 DNA to chromosome 19 of the transformed HEK293 cells. This indicates cytogenetic instability.​1​

Cell Morphology

In terms of cell morphology, it was long assumed that HEK293 were endothelial, epithelial or fibroblastic cells since those are abundant in embryonic kidneys. However, the presence of specific gene products and mRNA brought up speculations that they might have been neuronal cells. This could have been caused by adding the human adenovirus type 5. Consequently, the cell line should not serve as a representative in vitro model of classic kidney cells.

Sub Variants

The HEK293 lineage has several sub variants and derivatives, one of them being HEK 293T. What distinguishes the HEK 293T from its parent cell line is that the derivative contains the SV40 large T antigen. This makes it possible for the transfected plasmids to undergo episomal replication and thus produce a large amount of recombinant protein or retrovirus.

Applications of HEK293 cell lines

The application of HEK293 and its daughter cell lines in biotechnology and research studies is very common and versatile as outlined in the following:

• receptor signaling: HEK293 are used for heterologous expression of cell membrane receptors and ion channels as well as the investigation of signaling pathways of G protein coupled receptors.
• protein production: HEK293 produce recombinant proteins at a large scale and can be used for making biotherapeutic proteins and vaccines.
• cancer research: HEK293 are tumorigenic and suitable for testing effects of cancer treatment.
• hosts for experiments in gene expression: HEK293 cells are often used for transfection in a gene to assay the expressed protein, a method commonly applied in the research of the inducible RNA interference system.
• propagation of adenoviral vectors: In virology, HEK293 serve as an alternative to pathogens, since they are a less dangerous option.

How to handle HEK293 cells

As for their handling, HEK293 cells are considered as a comparatively low-maintenance line in cell biology and can be grown in culture ex vivo. Nevertheless, there are certain recommended cell culture conditions to be met such as a humidified incubator at 37°C supplemented with 5% CO2 as well as a high-glucose media diet to keep the cells alive. Other culturing media such as broad-spectrum antibiotics can help to prevent common bacterial infections.

However, it is important to acknowledge that these are known for causing harm to cells and results. The cell line grows rapidly and can thus be passaged every couple of days. However, if cultured over a certain time, the health of the cells degrades which can affect their reliability in experiments. The commercially available HEK293 transfection reagents and kids, for example through ATCC, add additional convenience to growing this cell line.

Advantages and limitations in using HEK293

There are several advantages as well as limitations that come with HEK293:

Advantages of HEK293 are

• rapid doubling time (about 36 hours)
• ease of culture
• can be grown as adherent or suspension regarding proliferation
• high reproducibility and general consistency
• produces large amounts of recombinant proteins, especially those within plasmid vectors containing the CMV promoter
• transient and stable gene expression
• ease of transfection through the use of physical and chemical methods

Limitations of HEK293 are

• danger of bacterial contamination potentially causing problems with gene expression and risking to lose cell viability
• particularly prone to viral contamination
• the length of its culture period negatively impacts the health of the cells

CHO cells — alternative to HEK293

CHO cells are a popular alternative to HEK293 cells. Due to their advantages, HEK293 cells are the second most frequently used line in cell biology and biotechnology right after HeLa cells. However, there are alternatives, such as the Chinese hamster ovary cells (CHO cells), able to overcome some of the above mentioned limitations. This mammalian cell line is especially popular due to its animal origin. This reduces the risk of propagation of human viruses and thereby minimizes production loss and increases biosafety. On top of this, CHO cells and HEK293 cells share many advantages, such as the robust growth and a high protein production, and are therefore commonly used in bio research and production of biotherapeutics, recombinant antibodies, and APIs. For evitria, CHO cells are thus the obvious choice for the expression of recombinant antibodies guaranteeing maximum quality in minimum time.