Explainer on 293 or HEK cells and their use in COVID vaccine research

A type of human cells used in research called HEK cells or 293s has been much in the news lately. This is mainly related to COVID-19 vaccine research. There seem to be many misunderstandings about 293 cells. For that reason, I thought I’d do this post to clear things up. I’ve been working with these cells for almost 3 decades so I know them well.

If you scroll down there’s also a video version of this post available.

What’s in this article?

What are cultured cells and how are they named? | What are HEK cells? | Why do we use 293s in research? | When I first used these cells | Clearing up confusion on 293 cells and COVID-19 | References

What are cultured cells and how are they named?

First, I’ll give a short introduction to cells grown in the lab more generally.

The cell lines that we use in research labs often go by basic alpha-numeric names. A cell line is generally considered immortal. This means we can grow it forever in the lab without it senescing. A common mouse fibroblast line is called 3T3s. Sometimes these names have an obscure source. It could be their place on an array of tissues culture wells or other lab details. It seems 3T3 cells were named after the protocol used to make them, “3-day transfer, inoculum 3×105 cells”. Other times they were named after patients. This was the case with HeLa cells, which came from Henrietta Lacks.

Many researchers also work with primary cells, which are generally defined as cells recently isolated. Primary cells can come from an organism like a person or a mouse.  Primary cells are mortal. We usually can only grow them for a matter of weeks. They functionally grow old quickly. The advantage of primary cells is that they are as close to normal as cells get in a lab. They generally have not had any major changes.

By contrast, cell lines can possibly acquire mutations over time. Such cells often were made into an immortal cell line in the first place by scientists. Researchers intentionally introducing cancer-related genes or something like TERT into them.

What are HEK cells?

The full name of 293 cells is HEK293 but they are sometimes called just HEK cells too. The acronym HEK reflects their identity as human embryonic kidney cells. Apparently, the 293 part of the name came simply from the experiment number from the scientist, postdoc Frank Graham, who isolated them while in Alex van der Eb’s lab.

These epithelial cells were derived by Graham way back in 1973 and since then have likely been used in hundreds of thousands of experiments, many that have been important for human health.

The first paper I could find describing 293 cells was from 1977. The paper describes how the primary embryonic kidney cells were immortalized by adding adenovirus DNA to them.

On another level, it’s interesting to read the paper, reflecting a different era in biomedical science. It’s like science history in a bottle.

The exact tissue source, other than being a human fetus, remains unclear. The specific fetus that was the source of 293 cells could have been either from a spontaneous miscarriage or an abortion.

293 cells come in many different subtypes. One of the most common versions is called 293T, which has been transformed by the oncogene SV40 large T antigen. There are also 293FT cells and other versions of 293s. Often these generally go under the umbrella of being called 293s.

Why do we use 293s in research?

What are the reasons to grow 293s for research? These cells grow quickly in culture, doubling in number more than once a day, which along with their ease of transfection (a term meaning introducing DNA into them often for protein or virus production) makes them a great lab tool.

While there are other methods to make proteins in cells including in bacterial cells, 293s are especially quick and powerful protein makers. And they produce human proteins with typical post-translational modifications like phosphorylation. Such modifications can be important and absent in proteins made in bacteria. Proteins made in human cells are also more likely to fold properly and be functional than those produced in bacteria.

In addition, 293s are really excellent at producing viruses for research. PLAT-A and PLAT-E cells, which are used to make viruses for research, were derived from 293s as well.

You can see the distinctive 293 cell appearance in culture in the lab in the image above from my own lab. The many round cells are in the process of dividing. 293s only attach to the cell culture plates relatively loosely, which can also give them a more rounded appearance than typical cells.

When I first used HEK cells in research

I remember the exact moment I heard about 293 cells for the first time as a scientist, which is kind of unusual as often such things are not clearly recalled.

I was a graduate student in the lab of Mark Kamps at UC San Diego. My work had mainly been with 3T3 cells and then previous to that as a technician in the Lane lab down the hall earlier on I had been working with human umbilical cord endothelial cells. Although I had overexpressed genes in 3T3 cells as a student before I had done this by transducing them with a virus encoding those genes.

A student from another lab brought up the idea of working with 293 cells because for another experiment we wanted to transfect cells and get high expression of the gene and protein in question.

It’s hard to believe that was more than 25 years ago! I was impressed at the time by how easy it was to transfect (meaning put DNA into cells) 293s and how much protein they crank out.

They are a valuable tool that my lab and thousands of others in academia and industry still use today.

Clearing up confusion on HEK cells and COVID-19

What’s the deal with 293 cells and COVID-19 vaccine production?

There has been a lot of confusion and even misinformation about 293 cells out there related to the pandemic.

Different types of 293 cells have been used in research on COVID-19 vaccines by different manufacturers. However, there are no human cells in the actual vaccines. Some have gotten that wrong.

Also, 293s are not stem cells and, more specifically, are quite different from human embryonic stem cells. This has also often been an area of confusion.

What about at the ethical or moral level? I personally see no problem in vaccine research and production using 293 or other human cells.

The uncertainty over the origin of the fetus used to make 293 cells leaves things a little fuzzy in that regard, but I’d said it also makes it harder to somehow condemn 293 cells as immoral to use in research, if that’s one’s agenda.

How are HEK cells used in COVID-19 vaccine research?

Vaccine production involves extensive research. For some of the reasons mentioned earlier, HEK cells are ideally suited for this research. For instance, the cells were instrumental in validating protein expression from the RNAs and folding in BNT 162b vaccine developmental research and for similar research for the Moderna vaccine. (HT to Brian GADD who pointed to this preprints on Twitter during a discussion there.)

Given the urgency of the pandemic, the speed and tractability of 293 cells was crucial and likely saved many lives by speeding effective vaccine development.

National Geographic has a nice broader explainer on the use of fetal cells in life sciences research that also mentions HEK cells too in relation to COVID vaccine production.

After already being used in the research reported in more than 50,000 publications, HEK 293 cells will continue to aid the life sciences and save lives in the future.

References

• Characteristics of a Human Cell Line Transformed by DNA from Human Adenovirus Type 5, Journal of General Virology, Graham, et al. 1977.
• Moral Guidance on Using COVID-19 Vaccines Developed with Human Fetal Cell Lines, The Journal of the Witherspoon Council.
• Human Embryonic Kidney 293 Cells: A Vehicle for Biopharmaceutical Manufacturing, Structural Biology, and Electrophysiology. Cells Tissues Organs.

• BNT162b vaccines are immunogenic and protect non-human primates against SARS-CoV-2, preprint

• SARS-CoV-2 mRNA Vaccine Development Enabled by Prototype Pathogen Preparedness, preprint

• Nat Geo explainer on the use of fetal cells in research.