Fetal bovine serum—a cell culture dilemma

Valk, Jan van der

doi:10.1126/science.abm1317

Cited by 39 publications

(35 citation statements)

References 13 publications

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“…Although alternatives are available, FBS is still the most commonly used culture medium supplement. However, its use is increasingly considered controversial due to several safety, scientific and ethical concerns ( 10 , 11 ). Also, for osteoclast-osteoblast co-cultures with human MCs and MSCs as progenitors, aiming at mimicking human bone remodeling and related pathologies in vitro¸ FBS is still the common standard ( 5 ).…”

Section: Discussionmentioning

confidence: 99%

“…Fetal bovine serum (FBS) is a culture medium supplement sourced from unborn calves at the slaughterhouse ( 10 ). FBS is currently easily available, relatively inexpensive, and it contains an excess of nutrients and proteins that support cell adhesion, growth, and proliferation.…”

Section: Introductionmentioning

confidence: 99%

“…FBS is currently easily available, relatively inexpensive, and it contains an excess of nutrients and proteins that support cell adhesion, growth, and proliferation. As a result, FBS is historically the most commonly used medium supplement for in vitro cultures, including for osteoclast-osteoblast co-cultures ( 5 , 10 ). However, several safety, scientific, and ethical concerns against the use of FBS have been raised ( 10 , 11 ).…”

Section: Introductionmentioning

confidence: 99%

“…As a result, FBS is historically the most commonly used medium supplement for in vitro cultures, including for osteoclast-osteoblast co-cultures ( 5 , 10 ). However, several safety, scientific, and ethical concerns against the use of FBS have been raised ( 10 , 11 ). Batch to batch variation, zoonotic pathogens, and xenogeneic proteins that are incompatible with human physiology may cause undesired and irreproducible experimental results ( 10 , 12 , 13 ).…”

Section: Introductionmentioning

confidence: 99%

“…However, several safety, scientific, and ethical concerns against the use of FBS have been raised ( 10 , 11 ). Batch to batch variation, zoonotic pathogens, and xenogeneic proteins that are incompatible with human physiology may cause undesired and irreproducible experimental results ( 10 , 12 , 13 ). In addition, with the aim to comply to the 3Rs, the use of animal components for in vitro alternatives to animal experiments is controversial ( 11 ).…”

Section: Introductionmentioning

confidence: 99%

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Human Platelet Lysate as Alternative of Fetal Bovine Serum for Enhanced Human In Vitro Bone Resorption and Remodeling

2022

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IntroductionTo study human physiological and pathological bone remodeling while addressing the principle of replacement, reduction and refinement of animal experiments (3Rs), human in vitro bone remodeling models are being developed. Despite increasing safety-, scientific-, and ethical concerns, fetal bovine serum (FBS), a nutritional medium supplement, is still routinely used in these models. To comply with the 3Rs and to improve the reproducibility of such in vitro models, xenogeneic-free medium supplements should be investigated. Human platelet lysate (hPL) might be a good alternative as it has been shown to accelerate osteogenic differentiation of mesenchymal stromal cells (MSCs) and improve subsequent mineralization. However, for a human in vitro bone model, hPL should also be able to adequately support osteoclastic differentiation and subsequent bone resorption. In addition, optimizing co-culture medium conditions in mono-cultures might lead to unequal stimulation of co-cultured cells.MethodsWe compared supplementation with 10% FBS vs. 10%, 5%, and 2.5% hPL for osteoclast formation and resorption by human monocytes (MCs) in mono-culture and in co-culture with (osteogenically stimulated) human MSCs.Results and DiscussionSupplementation of hPL can lead to a less donor-dependent and more homogeneous osteoclastic differentiation of MCs when compared to supplementation with 10% FBS. In co-cultures, osteoclastic differentiation and resorption in the 10% FBS group was almost completely inhibited by MSCs, while the supplementation with hPL still allowed for resorption, mostly at low concentrations. The addition of hPL to osteogenically stimulated MSC mono- and MC-MSC co-cultures resulted in osteogenic differentiation and bone-like matrix formation, mostly at high concentrations.ConclusionWe conclude that hPL could support both osteoclastic differentiation of human MCs and osteogenic differentiation of human MSCs in mono- and in co-culture, and that this can be balanced by the hPL concentration. Thus, the use of hPL could limit the need for FBS, which is currently commonly accepted for in vitro bone remodeling models.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Human Platelet Lysate as Alternative of Fetal Bovine Serum for Enhanced Human In Vitro Bone Resorption and Remodeling

2022

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Generation of stable suspension producer cell lines for serum‐free lentivirus production

Klimpel,

Terrao,

Bräuer

et al. 2024

Biotechnology Journal

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The production of lentiviral vectors (LVs) pseudotyped with the vesicular stomatitis virus envelope glycoprotein (VSV‐G) is limited by the associated cytotoxicity of the envelope and by the production methods used, such as transient transfection of adherent cell lines. In this study, we established stable suspension producer cell lines for scalable and serum‐free LV production derived from two stable, inducible packaging cell lines, named GPRG and GPRTG. The established polyclonal producer cell lines produce self‐inactivating (SIN) LVs carrying a WAS‐T2A‐GFP construct at an average infectious titer of up to 4.64 × 107 TU mL−1 in a semi‐perfusion process in a shake flask and can be generated in less than two months. The derived monoclonal cell lines are functionally stable in continuous culture and produce an average infectious titer of up to 9.38 × 107 TU mL−1 in a semi‐perfusion shake flask process. The producer clones are able to maintain a productivity of >1 × 107 TU mL−1 day−1 for up to 29 consecutive days in a non‐optimized 5 L stirred‐tank bioreactor perfusion process, representing a major milestone in the field of LV manufacturing. As the producer cell lines are based on an inducible Tet‐off expression system, the established process allows LV production in the absence of inducers such as antibiotics. The purified LVs efficiently transduce human CD34+ cells, reducing the LV quantities required for gene and cell therapy applications.

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Development of a perfusion process for continuous lentivirus production using stable suspension producer cell lines

Klimpel

Terrao

Ching

et al. 2023

Biotech & Bioengineering

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The large‐scale production of clinical‐grade lentiviral vectors (LVs) for gene therapy applications is a remaining challenge. The use of adherent cell lines and methods like transient transfection are cost‐intensive and hamper process scalability as well as reproducibility. This study describes the use of two suspension‐adapted stable packaging cell lines, called GPRGs and GPRTGs, for the development of a scalable and serum‐free LV production process. Both stable packaging cell lines are based on an inducible Tet‐off system, thus requiring doxycycline removal for initiation of the virus production. Therefore, we compared different methods for doxycycline removal and inoculated three independent 5 L bioreactors using a scalable induction method by dilution, an acoustic cell washer and manual centrifugation. The bioreactors were inoculated with a stable producer cell line encoding for a LV carrying a clinically relevant gene. LV production was performed in perfusion mode using a cell retention device based on acoustic wave separation. Comparable cell‐specific productivities were obtained with all three methods and cumulative functional yields up to 6.36 × 1011 transducing units per bioreactor were generated in a 234‐h long process, demonstrating the usability of stable Tet‐off cell lines for an easily scalable suspension process. Remarkably, cell viabilities >90% were maintained at high cell densities without compromising productivity throughout the whole process, allowing to further extend the process time. Given its low effects of toxicity during virus production, the presented cell lines are excellent candidates to develop a fully continuous LV production process to overcome the existing bottlenecks in LV manufacturing.

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Fetal bovine serum—a cell culture dilemma

Abstract: Ethical and possible reproducibility issues arise when using fetal bovine serum in cell culture media

Cited by 39 publications

References 13 publications

Human Platelet Lysate as Alternative of Fetal Bovine Serum for Enhanced Human In Vitro Bone Resorption and Remodeling

Human Platelet Lysate as Alternative of Fetal Bovine Serum for Enhanced Human In Vitro Bone Resorption and Remodeling

Generation of stable suspension producer cell lines for serum‐free lentivirus production

Development of a perfusion process for continuous lentivirus production using stable suspension producer cell lines

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