Alternatives to Fetal Bovine Serum in Cultivating Cultured Meat

An Academic Review

In recent times, the realm of lab-grown meat has captured global attention, making significant strides in its development and overcoming various challenges. Just a day ago, Bene Meat, a Czech firm, secured EU approval for its lab-grown pet food, following the approval for lab-grown meat sales in the US earlier this year. With all this action, it is safe to say that lab-grown meat is getting closer and closer to completely revolutionizing our food systems.

Now, as you can probably guess, lab-grown meat is meat grown without any animal involvement. Funnily enough, up until very recently, lab-grown meat did vitally depend on an animal-derived substance: Fetal Bovine Serum. In this review, I will be consolidating my learnings from researching on the latest advancements in the field, both from the lens of lab-grown meat as well as general cell culture.

Note: If you are not acquainted with the space, here are some suggested prior reading links.

Outline of Paper

1. Introduction
2. Fetal Bovine Media as Culture Media
   — 2.1: Fetal Bovine Serum as an Effective Culture Medium
   — 2.2: Ethical, Environmental and Scientific Considerations of FBS
3. General Alternatives to the Use Of FBS in media
   — 3.1: Bovine Ocular Fluid
   —3.2: Sericin
   — 3.3: Human Platelet Lysate (HPL)
   —3.4: Heat Inactivated Coelomic Fluid (HI-CF)
   — 3.5: HPL x HI-CF — A Potential Alternative Outlined By Researchers
4. Serum-Free Media for Use in Cultured Meat Production
   — 4.1: Cell types in meat
   — 4.2: Various Media Currently Researched for Use in Cultured Meat
   — — — 4.2.1: Beefy-9 — A Serum-free Culture Media for Expansion of Bovine Satellite Cells (BSCs)
   — — — 4.2.2: Replacing the rAlbumin in Beefy-9: Beefy-R
   — — — 4.2.1: Serum Free Differentiation Medium (SFDM) — Company Spotlight: Mosa Meat
5. Conclusion
6. References

Introduction

Presently, one of the key hurdles to the commercialization and mass adoption of cultured meat is its production costs. Obviously, customers would be less willing to pay more while they could get, in their opinion, the less “artificial” stuff for less.

Upon taking a closer look, we see that a significant cost driver for cultivated meat is the culture medium. In fact, 55% to >95% of marginal costs can be attributed to culture medium alone. The culture medium serves as the source of all growth factors, hormones, amino acids, vitamins, inorganic salts and macromolecules, replicating the in-vivo environment of the cell. It is the diffuse environment of the cell, which is an essential aspect of the cell microenvironment. It consists of a basal medium and is often supplemented by animal-derived serum. While a basal medium formulation (eg. DMEM, Ham’s F12 Nutrient Mixture , MCDB etc…) is often sufficient to keep cells alive for a short period of time, if we want to achieve the scale we are thinking of for cultured meat, there is a need for serum.

Fetal Bovine Media as Serum

In the case of cultured meat, the serum we use is fetal bovine serum (FBS). As you can probably imagine, it comes with a set of problems of its own across the scientific, moral and economic facets, but it does a neat job of providing everything that the cell needs in order to survive. Hence, for the culture of bovine satellite cells, we traditionally rely on fetal bovine serum despite the knowledge that it is unsustainable.

3.1: Fetal Bovine Serum as an Effective Supplement to Culture Medium

The aim of sera in general is to provide hormonal factors stimulating cell growth and proliferation, binding and transport proteins and stabilizing and detoxifying factors needed to maintain pH. Fetal bovine serum is essentially a smoothie of all the necessary growth factors, macro and micromolecules required for the proliferation, differentiation and maintenance of cells.

FBS, as the name suggests, is obtained from the fetus of slaughtered pregnant cows usually through cardiac puncture (though it can happen through puncture of umbilical vein or jugular vein as well). First, it is sterilized and the blood is extracted and stored to promote coagulation. With increased coagulation, the quality of serum increases.

The serum is separated from the blood corpuscles and cruor (clotting factors) via centrifugation. It is further sterilized and filtered by a variety of methods, including the following:

• 0.1 micrometer triple filtration
• Irradiation (via gamma rays)
• Removal of potential bacterial contamination

The obtained serum is further assessed for a variety of quality controls, including residual microbial/viral contamination, endotoxin content etc…Finally, it is frozen and packaged to keep the growth factors alive.

The reason why research scientists and tissue engineers love FBS so much is because of its many advantages in cell culture.

Firstly, FBS is rich in growth factors, which are integral to ensuring cell growth, metabolism and proliferation. Some of the growth factors present in FBS are cytokines, basic fibroblast growth factor (bFGF), endothelial cell growth factor (ECGF), interleukins, interferons and more. A full list of these factors is available here.

Some of these growth factors also affect differentiation, such as epidermal growth factor, insulin-like growth factor and so on, which can shorten the cell culture period, effectively reducing costs.

Saccharides and amino acids are also present in the sera and it support the energy needs of the cell for proliferation and metabolism.

Secondly, the presence of the protein albumin, a non-adhesive protein, in high amounts plays an integral role in cell adhesion. Unlike suspension cells, adherent cells such as the ones used in cultured meat require a substrate to grow on; they grow as monolayers, sticking to an extracellular matrix. Hence, the secure attachment of cells to culture dishes is an important factor when it comes to early cell growth. According to this paper, cell adhesion to the matrix is activated and promoted by the presence of low levels of adhesion proteins (eg. fibronectin) coupled with higher concentrations of non-adhesion proteins (eg. osteonectin). Here, albumin activates adhesion and spreading factors.

Albumin constitutes 60% of total proteins in the serum and is vital to making the medium what it is. It also acts as an antioxidant that stabilizes and transports compounds, increases availability and solubility of beneficial factors while reducing the accumulation of harmful byproducts. It non-specifically binds to poorly soluble substances, as a circulating protein.

Aside from albumin, it also contains 1800 proteins and over 4000 metabolites. As aforementioned, a list of the major components present in the serum can be found here.

Thirdly, FBS contains low levels of immunoglobulin and complement content. This is due to the developmentally immature immune systems present in fetal cows. This reduces immune response from antibodies. Moreover, FBS contains low levels of gamma-globulins (antibodies). Gamma globulins are proteins which are usually synthesized in order to eliminate an antigen. It does so by specifically binding to them and activating an immune response. These are notoriously known for inhibiting cell growth in culture.

Finally, it also helps improve the pH buffering capacity of the medium.

All of these factors contribute to the reputation of FBS as the perfect cocktail of growth factors and nutrients, the full package for optimal cell growth. It is also important to note that FBS is not just used in cultured meat, but is used for almost every cell culture due to its universal properties. Hence, by using FBS-supplemented media, we have a reduced need to spend time and effort developing a specific, optimized media formulation for each cell type. However, as we progress through research, many ethical, scientific and economic concerns are being raised about its usage.

3.2: Ethical, Environmental and Scientific Considerations of FBS

The fundamental idea that inspired cell-based meat is to elimination of any animal involvement. However, by depending on FBS for something as integral as the culture media, we are quite literally going against what we set out to do.

Aside from that, there are a few other concerns discussed under this. These can be categorized into three buckets:

1. Scientific
   * FBS is chemically undefined, leading to batch-to-batch variability: FBS contains a large number of components, the true composition of which is unknown. The composition varies based on diet of the cow, seasonality, quality and identity of the hormones received by the mother and gestational age of the fetus. Hence, quality control and testing to ensure quality become inconvenient from an economic perspectives, especially in the case of smaller academic labs. Moreover, this can add to external costs.
   *Risk of contamination: There is a potential risk of contamination from multiple organisms (eg. mycoplasma, viruses and bovine spongiform encephalopathy — Mad Cow Disease). While there are a variety of methods to filter/elimate these, there is no guarantee and some methods also damage growth factors and other proteins present in the serum. Moreover, it can potentially interfere with phenotypic cell stability and influence expression.
2. Economic:
   *Limited global Supply: It has been seen that while the demand for FBS as a supplement increases, there is relatively stagnant growth in its availability. This suggests that we have reached “peak serum”. This can also be accounted to a fault in the system; FBS being a byproduct of a more profitable industry, farmers are not incentivized to increase cattle herds to meet a future demand of FBS.
   *Cost (and it’s not getting better): The prices of FBS is ridiculously high and is one of the highest relative to other components of cell based meat. With the limited global supply, the prices of FBS has increase 3 fold over the past years.
   *Loosely regulated market: Soaring demands coupled with a market with little regulations set form the perfect environment for exploitation.
3. Ethical:
   *Animal Welfare: Roughly 2 million fetal calves are used in serum collection annually, totally approximately 800,000 liters of FBS produced per year (Good Food Institute). As aforementioned, the collection process usually happens via cardiac puncture, where a syringe is placed directly into the beating heart. This raises further ethical considerations, as many believe that the fetus can consciously feel pain in this scenario.

Due to these reasons, there is a greater push by leaders and experts to steer away from the use of FBS and to use more sustainable alternatives.

General Alternatives to the Use of Fetal Bovine Serum (FBS)

As of now, there are 260 different cell types that can be cultured using serum-free media. However, none of them have the universality that FBS offers. As of now, there is no single alternate that can replace FBS to culture all cell types. In this section, we will be discussing the many general alternatives to the use of FBS, not specifically in the field of cultured meat.

3.1: SR-2.05 — Bovine Ocular Fluid

Bovine ocular fluid is fluid collected from the eye of a cow within 6 hours after slaughter. The fluid is then surface sterilized and centrifuged, giving way to a clear supernatant, which was then collected, filtered and stored.

To prepare the serum replacement (SR-2.05), roughly 35% of sheep’s defibrinated plasma and 1.5% of serum albumin is combined with the bovine ocular fluid. This novel serum replacement is especially useful for the culture of human bone marrow fibroblasts, vero cells (cells extracted from the kidney of an African Green Monkey), WISH (human amniotic epithelial cells) cells and it is even seen that these cells grow better and quicker with this supplementation compared to FBS.

Note: the basal media used to test the serum replacement in all cell lines was Eagle’s medium (supplemented by 10% SR-2.05).

In the below figure, you can see the influence of SR-2.05 on the growth of chicken embryonic fibroblasts.

PD — population doublings, CPD — cumulative population doublings, GI — growth index | Source

Though the chemical composition is undefined, it contains fibronectin (a classical cell adhesion protein), hypoxanthine, insulin-like growth factor, vascular endothelial growth factor, etc… just like FBS.

However, it is not likely to be used in the cultured meat space due to two reasons:

• The effectiveness of the ocular-fluid based supplement needs to be studied for stem cells
• It may not be feasible to rely entirely on this medium due to its limited quantities.

Even if we were to confirm its efficacy in the cultured meat space, we could use it in certain stages. A particularly interesting feature of this medium is that practically no adaptation is needed to use it. For instance, the cells could be grown in the medium supplemented by FBS in the first passage, followed by another medium supplemented by SR-2.05 in the next without any problem.

3.2: Sericin — Silk Protein

Sericin is a type of globular glue protein present in silk produced by the silkmoth Bombyx mori. It mainly consists of polar amino acids like aspartic acid, serine. It is obtained by degumming from raw silk, giving you a shiny, semitransparent sheet of silk.

Sericin as a serum replacement offers many benefits. Firstly, it has been shown to boost cell attachment of human skin fibroblast and growth of mouse fibroblasts due to its mitogenic capacity (i.e. ability to induce cell division — mitosis). Secondly, sericin contains methionine and cysteine which enhance collagen production and has a positive effect on cell proliferation. Thirdly, sericin improves cell survival rates by decreasing lactate dehydrogenase activity — an important indicator of cell death. Finally, despite the inactivating/denaturing action of heat on proteins, sericin maintained mitogenic activity even after being exposed to heat for long periods of time (20–60 minutes).

As per this table, it is inferred that sericin has a positive effect on many different cell types including human/animal cells, hybridomas, tumour cells, oocytes, embryos, organs and even insect cells. In fact, in the case of Chinese hamster ovary cells (CHO), African green monkey kidney cells and HeLa cells, there was virtually no difference between FBS and sericin. This is a promising start.

However, while sericin as a supplement has been extensively studied as an additive in cell culture, a quick search on Google Scholar reveals that there hasn’t been any specific studies of using this replacement for cultured meat specifically.

3.3: Human Platelet Lysate (HPL)

As we know, in our body, when platelets are activated, a wound that may once have been bleeding recovers.

In our body, platelets play a key role in repair. They are activated in the presence of damaged tissue (when they come in contact with connective tissue etc), delivering many types of growth factors, including platelet-derived growth factor (PDGF), transforming growth factor-beta (TGFß), basic fibroblast growth factor (bFGF), fibronectin, P-selectin and more. They perform this delivery by a method known as exocytosis, where the molecules are expelled out to the cell exterior.

With all the hard work it does in our body, imagine its potential to be used in cell culture!

This is exactly what’s being done — outdated human donor platelets are obtained by different methods, which undergo centrifugation (so as to remove the storage solution — platelet additive solution (PAS)). It is then washed with 0.9% saline solution and resuspended in the same. This suspension is stored at -20 C and finally, our serum replacement additive is prepared by three freezing/thawing cycles (for most efficient platelet activation).

HPL has strong mitogens, promoting proliferation and attachment of cells. Another advantage of the use of HPL is that it supports both suspension and adherent cells. However, it is mainly used in the medical space to bring about innovation and advancements such as regenerative medicine. Hence, it is not as accessible to use at a larger scale.

3.4: Earthworm Heat Inactivated Coelomic Fluid (HI-CF)

Coelomic fluid (CF) is the fluid that circulates through the earthworm’s body segments, acting as a hydroskeleton as well as facilitating the transport of immune cells like coelomocytes and nutrients.

CF is derived from the mesenchymal lining, consisting of a watery matrix and the plasma. It has been shown to contain various bioactive compounds such as proteins and exhibit a variety of biological functions such as antibacterial, anticancer, hemagglutinating and proteolytic activities. Earthworms contain a large number of metabolites and growth factors, making it ideal to serve as a serum replacement for FBS. In addition to that, coelomic fluids of earthworms, Eudrilus eugeniae, Eisenia spp. and Lumbricus spp. are rich in riboflavin, which has been shown to promote proliferation and proliferative potential of certain cells. Furthermore, Flavin Adenine Dinucleotide (FAD), a product of riboflavin, promotes the neuronal differentiation of human neural stem cells (from fetal brain as well as iPSCs). It has also been shown to have the mitogenic effect, possibly from the presence of the factor CMF (coelomic mitogenic factor). Finally, earthworms being lower invertebrates have no immunoglobulin content, making it even more favourable.

The cells used in cultured meat are adherent cells, which means that they grow attached to a surface. However, in HI-CF, the presence of fibrinolytic enzymes hinders the attachment of these cells. Moreover, this medium lacks the cell adherent factors such as fibronectin. This means that for initial attachment, the medium would require cell adherent factors. Hence, this too does not favour the growth of BSCs to the same degree that FBS does.

3.5: HPL x HI-CF — A Potential Alternative Outlined By Researchers

The authors of this paper have outlined a potential alternative to FBS by combining HI-CF obtained from the earthworm P. Excavatus (due to its antibacterial properties, and wide availability) and HPL in a ratio of 1:9 or 2:8 and supplementing it in basal medium.

In this case, the primary function of HPL is to support the attachment and growth of cells (which HI-CF cannot do independently). Unlike HI-CF, HPL contains attachment and spreading factors such as P-selectin, Von willebrand Factor, fibronectin etc… The below figure illustrates this alternative in detail.

Source

Serum-Free Media for Use in Cultured Meat production

In this section, we will be discussing serum-free media for use in cultured meat production specifically. As you may have gathered by now, most media are designed for specific cell types. In the case of cultured meat, we deal with a variety of cell types which have to fuse together. In order to propose alternate media, we need to know what types of cell we are trying to culture. The next section outlines the various cell types seen in meat:

4.1: Composition of Meat & Cells Used in Cell-Based Meat

Meat is comprised of roughly 90% muscle fibres, 10% fat and connective tissue and less than 1% blood. The key cell type present is the skeletal myocyte (skeletal muscle cells), but the following are also significant cell types that play a key role in making meat, meat:

• Adipocytes: fat cells — store energy as fat
• Fibroblasts: contributes to formation and maintenance of structural framework of tissues
• Chrondrocytes: cells that make up the cartilage and sustain the ECM
• Hematopoietic Stem Cells: multipotent stem cells that give rise to various types of blood cells

As we are culturing these cells, we use stem cells in the case of cultivated meat. By choosing to use stem cells, we open ourselves to two possibilities: Adult stem cells and pluripotent stem cells. You can learn more about these types from this article, written by yours truly ;)

Anyways, both of these come with their own set of pros and cons. For instance, when dealing with adult stem cells, we have to deal with the limitation of reduced proliferative capacity and maintenance relative to pluripotent stem cells.

In the case of pluripotent stem cells, more specifically iPSCs, there is more research needed before we introduce these cells in the cell culture process — especially when it comes to cell reprogramming. On the other hand, embryonic stem cells are difficult to harvest. These stem cells are derived from the inner cell mass of a blastocyst. Finally, even if we were to obtain these stem cells, there are no well-established differentiation protocols, which just ends up being more work.

4.2: Various Media Currently Researched for Use in Cultured Meat

While the media that we discussed earlier are potential alternatives to FBS, they haven’t been studied or researched in-depth for their use in cultured meat. Some of the popular ones studied include:

4.2.1: Beefy-9 — A Serum-free Culture Media for Expansion of Bovine Satellite Cells (BSCs)

Beefy-9 is a B8-inspired serum-free media for culturing bovine satelline cells. B8 is a serum-free formulation designed for the culturing of human pluripotent stem cells. It is improved and adapted to the use for culturing BSCs by adding a protein, albumin. This resultant media has been appropriately named as “Beefy-9”.

While B8 media supplementation can lower the need for serum, it still does not completely eliminate it. In fact, in the same study, it is shown that a serum reduction of 87.5% did not impact cell growth significantly. However, using B8 alone was not sustainable and cell growth did not continue for long after elimination of serum.

In order to completely eliminate the use of serum, the media was tested with the addition of a variety of supplements (eg. interleukin-6, curcumin, PDGF-BB etc…) in different concentrations. The growth of the cells were then analyzed over the duration of 4 days. From this, the most effective supplement was shown to be rAlbumin at the concentration of 800 μg/mL, leading to a 4x improvement in growth. Another possible combination that has been outlined in the paper is Beefy-9+ which is essentially Beefy-9 with 3.2mg/L of rAlbumin, said to have the best cell growth to media cost ratio

testing B8 with various supplements (Source)

However, recombinant albumin (rAlbumin) hindered cell attachment in passaging and so, it was found that cells had to be passaged in its absence. Moreover, a coating of a cell adhesive peptide such as 1.5 μg/cm2 of Vtn-N is required. The entire process has been outlined below.

passaging with Beefy-9 (Source)

Beefy-9 has proven effective in maintaining cell myogenicity and satellite cell identity, comparable to serum-containing media. An analysis of Pax7, an early satellite cell marker of skeletal cell differentiation, revealed that over 96% of cells in both serum-containing and Beefy-9-supplemented samples stained positive for Pax7, indicating no significant difference in Pax7 gene expression.

An interesting point to note here is that cells cultured in serum-free conditions accumulate long-term lipid droplets over long term culture. The authors of the paper hypothesize that this is due to the development of insulin resistance in cells due to the increased amount of insulin present in Beefy-9. However, lipids generally have a positive impact on meat flavour and so this can also be viewed as an advantage.

Additionally, the amount and composition of Beefy-9 can be altered, which presents several cost-saving opportunities, making it a feasible alternative to FBS. Even so, rAlbumin in Beefy-9, despite its many advantages, contributes to a significant percentage of total costs, rendering it economically unviable for large-scale utilization. In fact, depending on its concentration, the addition of albumin can result in a 50% to 400% cost increase.

4.2.2: Replacing the rAlbumin in Beefy-9: Beefy-R

As a replacement of rAlbumin in Beefy-9, oilseed protein isolates — more specifically, rapeseed protein isolate (RPI) — has been used to create what is known as “Beefy-R” culture medium. This medium is constituted by B8 with 0.4mg/mL RPI. This modified medium promotes relatively enhanced cell growth while maintaining the phenotype and differentiation capacity of BSCs.

Using oilseed protein isolates is especially beneficial due to its lower costs, scalability as well as natural abundance. It also produces results comparable to Beefy-9, as you’ll read.

The RPI has been obtained via the methods of alkali extraction, isoelectric precipitation, centrifugation and filtration, giving a reddish brown isolate. It has a relatively higher presence of proteins involved in cellular and metabolic processes, containing more binding and catalytic proteins. Moreover, in comparison with other OPIs, RPI has the highest amount of albumins and lowest amount of globulins, which definitely contributes to its efficacy as a culture medium. It also shows a relatively high abundance of oleosins, which are a type of stabilizing proteins in oil seeds.

Out of all oilseed protein isolates, RPI at optimal concentration (0.4 mg/mL) performed the best and was comparable in efficacy to albumin. As the graphs show, after a certain threshold concentration, the isolates seem to have an inhibitory effect on the growth of BSCs.

Efficacy of different OPIs | Source

Passaging was done via the same process as was used earlier (i.e. coating with vitronectin and withholding the rAlbumin/RPI while seeding till attachment), but it was observed that Beefy-R had improved growth compared to Beefy-9 over 4 passages. In fact, 2x as many cells grew over 2 weeks in Beefy-R (11.7 population doublings). However, this still hasn’t reached FBS level growth: 14.8 doublings.

population doublings in passaging (source)

Similar to Beefy-9, Beefy R also maintains satellite cell identity (shown as 99% of cells were Pax7 positive), myogenic capacity and presents enhanced differentiation of cells. It also reduced the abnormal extent of lipid accumulation observed in Beefy-9.

One downside to this medium is that there is lot-by-lot variability in the efficacy of RPI, which means that the concentration used would also need to change.

4.2.3: Serum Free Differentiation Medium (SFDM) — Company Spotlight: Mosa Meat

The Dutch startup Mosa Meat has overcome the hurdle of FBS to achieve muscle differentiation, which is a huge step into commercialization. This startup used transcriptomics and proteomics to develop a serum-free medium formulation that is able to induce differentiation in bovine satellite stem cells.

As of now, differentiation is induced via serum starvation (i.e. suddenly reducing the serum concentration present in the growth medium) from 20% concentration in growth medium to 2% concentration in differentiation medium. Researchers at Mosa Meat took a different approach to this by investigating the the changes induced by serum starvation on the transcriptome, which is essentially a complete set of RNA transcripts, of BSCs, to bring about the process of myogenic differentiation. This was done via RNA sequencing.

It was found that during serum starvation, the canonical differentiation markers (genes/proteins that indicate that differentiation has taken place) and myogenic transcription factors were consistently upregulated, i.e. increased in expression throughout the differentiation process. Some specific findings include:

• The myogenic regulator MYF5 was most highly expressed 24 hours following serum starvation
• Pax7 increased during first 24 hours and then downregulated (i.e. decreased in expression)
• There was increased expression of skeletal-muscle-specific myosins and myoblast fusogens, suggesting ongoing maturation of the newly differentiated cells
• At the same time, there was a decrease in expression of satellite cell markers, cell-adhesion-related and cell-cycle maintenance genes, which denote a shift in gene expression from satellite cells to mature muscle cells.

Moreover, surface receptors, which are certain proteins on the cell surface involved in signalling/receiving signals, too upregulated during serum starvation. The specific surface receptors were identified: IGF1R and IGF2R, which mediate signaling of insulin, IGF1, and IGF2, as well as receptors for transferrin (TFRC), lysophosphatidic acid (LPA; LPAR1), oxytocin (OXTR), glucagon (GCGR), and acetylcholine (CHRNA). Upregulated genes primarily encoded for proteins related to muscle development and function, protein folding and cell cycle inhibitors. Furthermore, the downregulated genes were associated with cell cycle regulation, which indicated that serum starvation induces differentiation through cell cycle arrest (preventing the cell from further divisions.

Researchers from Mosa Meat aimed to induce differentiation using serum free media (absence of serum = absence of serum starvation). They did so by introducing activatory ligands, which are molecules that bind to surface receptors and activate them, to the previously upregulated surface receptors. So, they supplemented basal media DMEM, with NaHCO3, -ascorbic acid 2-phosphate, EGF1, MEM amino acids and serum albumin to form serum-free base (SFB). SFB was then supplemented with insulin and transferrin and other activatory ligands such as glucagon and lysophosphatidic acid (LPA), which led to an increased fusion index of 38.1% — 41% (depending on concentration of LPA added), as compared to 39% via serum starvation. SFB, supplemented with transferrin, insulin and LPA, is known as serum-free differentiation medium.

**An important means to understand or “quantify” differentiation is by looking at the fusion index. Here, while SFB alone did not have any relevant impact on the fusion index, with supplementation, it was comparable to the index of serum starvation.

“Representative fluorescence images of SCs after 72h in SFB+T+I with additional supplementation of LPA, oxytocin, glucagon and acetylcholine (ACh). Green, desmin; blue, Hoechst. Scale bar, 500µm”

The transcriptomic changes were profiled, and it was seen that the overall changes were very similar, indicating that the same underlying biological mechanisms are being activated.

In this way, Mosa Meat has revolutionized cultured meat production by pioneering a first-principles-based approach to formulate a serum-free growth medium, setting the stage for a paradigm shift in the future of sustainable and cruelty-free meat cultivation. More details can be found in their paper, which this review references.

Conclusion

As the world starts welcoming the prospects of lab-grown meat as a potential substitute for meat, it is important to develop a low-cost animal free media formulation for cultured meat. While FBS is a very effective supplement for culture media, it comes with significant drawbacks in terms of the environmental, ethical and economical fronts.

Some of the more general alternatives to the use of FBS include potential supplementations such as bovine ocular fluid, sericin, human platelet lysate and HI-CF (and even a combination of HPL and HI-CF). It has been noted that most of them have not been studied for their use in culturing bovine satellite cells specifically and they come with their own limitations. Further study is needed especially to gauge the feasibility of sericin supplementation in basal media for the proliferation of BCSs as well as the combination of HI-CF and HPL.

In terms of specific alternatives to FBS in the cultured meat space, Beefy-R media seems to be the most promising, due to its low cost and comparable performance. Mosa Meat too has succeeded in creating their serum-free differentiation medium by harnessing transcriptomics and proteomics. In fact, Mosa Meat was the first cultivated meat company to gain B-Corp certification.

Sources & References

All the sources and references used to write this review have been linked in the below spreadsheet. I would also like to express my deepest gratitude towards Dr Andrew J Stout and Dr Tobias Messmer for sending me their papers for my reference as I wrote this review — I truly appreciate it!

https://docs.google.com/spreadsheets/d/15mMgML_C6NBMhKSlaOQclH9aF3cAclCWTijoXVXDzOM/edit?usp=sharing