How might FBS be affecting the behaviour of the cells you culture and potential implications for cell therapies

If you’re culturing cells in fetal bovine serum, odds are your human cells might now be expressing cow sugars on their surface.

Neu5Gc (N-glycolylneuraminic acid) is a sialic acid found on the cell surfaces of most mammals, including cows. Humans lost the ability to make it roughly 3 million years ago, thanks to a mutation in the CMAH gene, so human cells evolved a different sialic-acid landscape.

However, when you culture cells in FBS-supplemented media, they can take up Neu5Gc from serum. It’s then metabolically incorporated into the cell's own glycoproteins and glycolipids. And just like that, you have human cells displaying non-human sugars on their surfaces.

As you can probably imagine, this becomes a substantial problem when we start to think about cell therapies. Many people carry anti-Neu5Gc antibodies. Cells that have incorporated Neu5Gc during culture can become targets for that immune response. This is just one of many reasons why animal-derived culture components are given serious scrutiny when it comes to culture of cells going anywhere near a clinic.

For routine cell culture, this could serve as an unreported confound in glycan-based surface marker characterization, but I mostly call this out as a small illustration of the bigger picture.

The boundary between "the cell" and "the culture medium" is fuzzy - much, much more so than we like to admit. When you culture human cells in animal-derived serum, you're giving them molecules from another species; some of those molecules can be metabolised and become part of the cells themselves. Neu5Gc is only one example.

Of course Neu5Gc content isn't typically quantified as part of FBS lot to lot QC, so this is also one more dimension of "batch variability" that doesn’t make it into the methods section.

But ultimately, this is why regulators are placing greater emphasis on the control and qualification of animal-derived materials used in cell and gene therapy manufacturing. As more of these therapies move from the lab to patients, the pressure to use “animal-free” media and other inputs wherever possible will increase. And so that’s why a slow, but steady shift has been occurring. After all, why would you build a clinical manufacturing process around an animal-derived raw material - and take on the additional qualification burden that comes with it - if you can avoid it?

As always, literature below if you're keen to dive deeper!
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More on Neu5Gc specifically, and human cells: https://pubmed.ncbi.nlm.nih.gov/22692205/, https://pmc.ncbi.nlm.nih.gov/articles/PMC7174778/

More on Neu5Gc specifically, and human cells: https://pubmed.ncbi.nlm.nih.gov/22692205/, https://pmc.ncbi.nlm.nih.gov/articles/PMC7174778/

More on Neu5Gc specifically, and human cells: https://pubmed.ncbi.nlm.nih.gov/22692205/, https://pmc.ncbi.nlm.nih.gov/articles/PMC7174778/

More on the patient immune response to FBS-cultured cell therapies: bovine apolipoprotein B-100 https://pmc.ncbi.nlm.nih.gov/articles/PMC1924480/
More on the patient immune response to FBS-cultured cell therapies: bovine apolipoprotein B-100 https://pmc.ncbi.nlm.nih.gov/articles/PMC1924480/More on Neu5Gc specifically, and human cells: https://pubmed.ncbi.nlm.nih.gov/22692205/, https://pmc.ncbi.nlm.nih.gov/articles/PMC7174778/

More on the patient immune response to FBS-cultured cell therapies: bovine apolipoprotein B-100 https://pmc.ncbi.nlm.nih.gov/articles/PMC1924480/