Ethics
Antibodies are indispensable tools in research, being widely used in antigen-binding assays such as Western Blot, ELISA, Immunoprecipitation, and Flow Cytometry due to their high specificity and binding affinity to target molecules. However, precisely because they are considered so crucial and irreplaceable, the ethically complex and resource-intensive manufacturing processes behind the production of novel antibodies are often overlooked by those who have the privilege to access them.
It is important to examine how the commercially available antibodies are produced in order to understand the flaws of current systems and why alternative approaches are necessary. First, we must differentiate between monoclonal and polyclonal antibodies, as they diverge both in their production and applications for end-users.
Custom polyclonal Antibodies (pAbs)
Polyclonal Antibodies originate from multiple B-cell clones and recognize several epitopes on a single target antigen. For their production, animals such as rabbits, guinea pigs, rats, mice, goats, or chickens are immunized with a given antigen for several weeks. The resulting serum contains a heterogenous mixture of immunoglobulins. After the immunization protocol is completed, samples are sent to end-users, who can decide whether to continue immunization by extending the protocol, collect terminal bleeds, or terminate the project without further action. As the production of polyclonal antibodies results in a finite supply, larger animals are preferred, as they yield greater serum volume [2].
Custom Monoclonal Antibodies (mAbs)
The production of Monoclonal Antibodies, in contrast, derives from a single B-cell and exhibits remarkable specificity by binding to one specific epitope on the target antigen. Animals such as mice, rabbits, rats, hamsters, chickens, guinea pigs or goats are immunized through a series of injections over several weeks, and test bleeds are performed in order to assess immune response. The best responding animals are further boosted twice and their cells are then harvested for hybridoma fusion. In this step, antibody-producing B cells are fused with immortalized myeloma cells in vitro, creating a cell line able to produce antibodies indefinitely. This process takes around 6 months to be completed [1].
As raised by the EU Reference Laboratory for alternatives to animal testing (EURL ECVAM) Recommendation on Non-Animal-Derived antibodies, the current production of both monoclonal and polyclonal antibodies, still heavily relies on animal immunization, and is associated with sophisticated laboratory infrastructure and considerable costs [3]. This situation led us to rethink antibody technologies and develop alternatives guided by three core values: Animal Welfare, Decentralized Bioeconomy and Open Science.
OUR 3 CORE VALUES
Animal welfare
Hover here!Animal welfare
Despite the emergence of promising in vitro Technologies, there is a striking contrast between guidelines and reality. The EU Directive 2010/63/EU on the protection of animals used for scientific purposes states that animal-based methods and technologies should not be employed when non-animal alternatives capable of producing equivalent results are available [4]. However, fully animal-free alternatives for certain applications, such as antibody production, are not yet available to replace animal-based methods. Consequently, according to the Deutsches Zentrum zum Schutz von Versuchstieren (Bf3R) (German centre for the protection of laboratory animals), 35930 animals were used in monoclonal and polyclonal antibody production in Germany in 2023 alone [5].
To reflect on this issue, we have reached out to Jana Wilken, spokesperson for Tierversuche Verstehen (Understanding Animal Testing). She provided us with statistical numbers and emphasized that the debate on animal testing goes far beyond the traditional Three Rs (Replacement, Reduction, and Refinement). Transparency is crucial, as animals used in science are often forgotten and remain invisible to end-users who simply purchase reagents. She further made us reflect that while alternatives must always be pursued, scientists should also make sure that, when animals are used, their dignity and welfare is safeguarded.
More about Jana Wilken
Jana Wilken is a science communicator and spokesperson for Tierversuche Verstehen, an organization that provides transparent and accessible information about the use of animals in research in Germany. Through her work, she seeks to build dialogue between researchers and society, promote efforts to reduce animal use, and ensure that those animals essential to research are treated with fairness and care.
Bioeconomy
Hover here!Bioeconomy
The production of antibodies is currently highly centralized and relies heavily on large companies with specialized infrastructure. This dependency limits the autonomy of individual researchers and institutions, as they cannot produce novel antibodies themselves and must instead rely on ordering them from external providers. In conversation with Prof. Dr. Joachin Boldt, it became clear that we must always think about the challenges in biology on a global scale, particularly through the bioethical principle of Global Justice. As described by Dr. Joseph Millium and Dr. Ezekiel J. Emanuel in their book Global Justice and Bioethics, fairness and equity in distributing the benefits and burdens in biological research are essential [6]. The centralization of antibody production in companies reinforces the inequality in the scientific landscape, as only laboratories with sufficient investment can afford to order the needed antibodies. Since discovery in research is linked to the available technologies, the centralization of resources feeds a cycle of inequality that limits global scientific practice.
More about Joachim Boldt
Joachim Boldt is a bioethicist at the University of Freiburg whose work explores the ethical dimensions of synthetic biology. He invited us to reflect about the values of bioethics, especially Global Justice, and had a major influence on our project.
Open Science
Hover here!Open Science
To explore the value of Open Science, we spoke with Sebastian Cocioba, a strong advocate of democratizing knowledge through the sharing of tools, protocols, and transparent communication. Thus, it became clear to us that Open Science goes beyond making tools accessible, but also about promoting collaboration. Guided by this principle, we envisioned the outcome of our Project as a toolkit that would allow researchers to create custom binding molecules for their protein of interest directly in their own lab without depending on large companies or even on us, as developers. However, we recognize that the independence we aim to provide also entails the responsibility of ensuring safe and ethical application of our toolkit by end-users. The considerations surrounding potential risks and oversight are discussed further on our Safety page.
More about Sebastian S. Cocioba
Sebastian S. Cocioba describes himself as an independent researcher and a self-taught amateur biologist focused on plant biotechnology. He is deeply committed to democratizing scientific knowledge by contributing to the creation of open-source tools and protocols. He runs Binomica Labs, a non-profit focused on collaborative science, aiming to lower the barriers to entry for aspiring researchers.
Conclusion
It is safe to state that the use of custom antibodies in research represents both a fundamental tool and a persistent ethical and practical problem, as their production is deeply tied to animal use and remains unevenly distributed to researchers worldwide due to high costs and dependency on centralized companies. Our project aims to change this reality. A toolkit that would empower researchers to create custom binding molecules for their protein of interest directly in their own lab, democratizing access to essential tools and promoting innovation.
Integrated Human Practices
Introduction
Science thrives through collaboration - it is built by a community, not in isolation.
Every new discovery is shaped not only by data and experiments, but also by the people who question, challenge, and inspire us. Feedback and criticism are fundamental to the scientific process. They allow us to broaden our vision and identify blindspots, therefore improving our work and that of the entire scientific community.
A central goal of our project is to make science more accessible, and we see our toolkit as one step toward that vision. However, this can only be realized through ongoing dialogue and open communication. Integrating the perspectives of a diverse group of people, ranging from researchers in well-equipped labs to those working with limited resources has guided us to understand how to design a toolkit that is not only scientifically innovative, but is also able to provide help where it is needed. Integrated Human Practices is not a single step in our workflow, it is the continuous dialogue that drives us forward. Every discussion, whether with researchers, professors, non-profit organizations or broader communities, opens up new questions and possibilities.
To show how our project has been shaped by dialogue and diverse perspectives, we have created a visual map of our Integrated Human Practices. Think of it as a river: it begins with the broad flow of ideas and feedback, which then branches into different streams, each representing a distinct part of our project. These streams carry insights, reflections, and adjustments, ultimately merging to become CONCAVE, our project.
Outlook
Throughout our Integrated Human Practices journey, each conversation has broadened our perspective and revealed new directions for our project CONCAVE. The insights we gained highlight not only the versatility of Repebodies but also the importance of interdisciplinary collaboration for realizing their full potential.
Therapeutics: Future research could explore their use as receptors, clotting agents, or tumor-detecting molecules. Their small size, high specificity, and cost-effectiveness make them promising candidates for accessible medical solutions. To advance Repebodies toward clinical use would require steps comparable to antibody development, such as humanization, large-scale production testing, and immunogenicity screening. Understanding these processes helps define what further optimization is needed before Repebodies could become viable diagnostic or therapeutic tools.
Artificial Intelligence: Another promising avenue lies in the integration of artificial intelligence. AI-based design could greatly accelerate the generation of Repebody variants by predicting viable sequences based on structural models rather than relying solely on random mutagenesis. This would allow for a more targeted and efficient exploration of sequences, potentially improving the quality and expanding the diversity of our protein binders.
Patenting: Beyond the scientific aspects, we also considered the ethical and legal dimensions of our work. The question of whether to patent our project led us to reflect on the balance between innovation and openness. Ultimately, we decided to make our findings publicly available, reinforcing our commitment to accessibility and collaboration in science.
Looking ahead, these insights provide a clear path forward:
Developing Repebodies with improved biocompatibility, integrating computational design methods, and maintaining an open, ethical approach to innovation. Our Integrated Human Practices have shown us that progress emerges through dialogue - each new perspective brings us closer to making science more inclusive, adaptable, and impactful.
References
[1] Thermo Fisher Scientific. (n.d.). Custom monoclonal antibody production. Thermo Fisher Scientific. https://www.thermofisher.com/de/de/home/life-science/antibodies/custom-antibodies/custom-antibody-production/custom-monoclonal-antibody-production.html, retrieved on October 6, 2025
[2] Thermo Fisher Scientific. (n.d.). Custom polyclonal antibody production. Thermo Fisher Scientific. https://www.thermofisher.com/de/de/home/life-science/antibodies/custom-antibodies/custom-antibody-production/custom-polyclonal-antibody-production.html, retrieved on October 6, 2025
[3] Barroso, J., Halder, M., & Whelan, M. (2020). EURL ECVAM recommendation on non-animal-derived antibodies (EUR 30185 EN). Publications Office of the European Union. https://doi.org/10.2760/80554
[4] European Parliament & Council of the European Union. (2010, September 22). Directive 2010/63/EU on the protection of animals used for scientific purposes (OJ L 276, pp. 33–79). https://eur-lex.europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2010:276:0033:0079:en:PDF
[5] Deutsches Zentrum zum Schutz von Versuchstieren (Bf3R). (2023). Zahlen 2023 – Tabelle 14: Anzahl der Tiere in der Routineproduktion. In Versuchstierzahlen 2023. https://www.bf3r.de/angebote/versuchstierzahlen/versuchstierzahlen-2023/
[6] Millum, J., & Emanuel, E. J. (Eds.). (2012). A Global justice and bioethics. Oxford University Press.
The Signalling Factory had recently begun researching DARPins independently of us, and since our PI Nicole Gensch is a leading member of that research group, she approached us about the possibility of using DARPins in our project instead of Repebodies.
It would be most advantageous for us to begin with our work using Repebodies rather than DARPins, but if they prove to be difficult to handle, we would still be able to switch to DARPins. DARPins are generally more researched than Repebodies, which might be easier to use in a project of our scale, but also made our work less original.
Dr. Nicole Gensch informed us about possible synergies between our team working with the Signalling Factory with DARPins, such as genetic material exchange and shared knowledge. When asked about what advantages or disadvantages DARPins may have over Repebodies, we discussed the flexibility of the proteins, as well as what kinds of proteins they can bind. DARPins, compared to Repebodies, are much less flexible, which may cause the binding strength to decrease. However, DARPins are known to have incredibly high binding strengths against many proteins, including when compared to Repebodies, but this may be due to a lack of research on Repebodies. Additionally, the consensus sequence of DARPins is different and less compatible with the retron system in comparison to Repebodies. Dr. Nicole Gensch also discussed current research around DARPins with us. In general, it can be said that DARPins have been studied more than Repebodies, suggesting that choosing to work with Repebodies would make our project more novel. Furthermore, the combination of targeted protein degradation together with a binding domain was already done in DARPins with bioPROTAC. We also asked Dr. Nicole Gensch if DARPins are more soluble than Repebodies, or if they needed a similar strain to Origami to efficiently express them in bacteria. She responded that since DARPins do not contain any disulfide bridges, they should be easily expressible in the bacterial cytosol. However, she also recommended that we try expressing Repebodies in BL21, since they may still fold properly even without the knock-out mutations of Origami.
We decided to continue to work with Repebodies for the time being. Concurrently, we kept in mind that we could switch to DARPins if Repebodies proved to be too difficult to work with. We preferred Repebodies, since they would allow us to conduct more original research, and because they seemed to be more promising when integrated into the retron system we had planned.
In the initial phase of defining our project, we contacted Prof. Dr. Manfred Jung, an expert in chemical epigenetics, since his research group's focus involves inhibitor synthesis and assay development. We reached out to him to gain a broader understanding of Targeted Protein Degradation (TPD) technologies and discuss potential project ideas involving chimeric molecules designed to degrade aggregated proteins in the context of neurodegenerative diseases.
Development of a new targeting chimeric molecule for degrading protein aggregations involved in disease is not feasible in the scope of an iGEM project. However, we could explore different TPD approaches that do not require synthesis of a new chimeric molecule. We should contact Prof. Dr. Claudine Kraft, an autophagy expert.
Prof. Dr. Manfred Jung explained that TPD technologies are based on chimeric molecules specifically designed to target a protein of interest (POI) and induce its degradation. To develop a new targeting chimera, we would need to focus on a known protein with a well-characterized ligand and binding site. Designing and synthesizing a molecule is very complicated as it requires special equipment as well as expertise in handling it. In Prof. Jung's experience, it can take months to find a proper linker with an optimal length and address the stability issues and activity. Creating a new targeting chimera for a protein involved in disease is therefore not feasible as an iGEM project. However, we could explore alternative TPD technologies that do not require novel chemical synthesis but instead involve tagging the POI with a degradation tag like HaloTAG or dTAG. This approach could serve as a proof of concept and be linked with autophagy as a novel application. However, he explained these technologies could be less efficient because the POI is not directly marked for degradation. The POI gets fused to the TAG and then the TAG is degraded, along with the POI. We further asked him about AutoTAC, a TPD technology that was used to degrade alpha-Synuclein in Parkinson's disease on mice. Prof. Dr. Manfred Jung mentioned that such approaches are often being commercialized by various companies and advised investigating related patents and further publications. He recommended that we reach out to Prof. Dr. Claudine Kraft, who specializes in autophagy research.
We discarded the idea of synthesizing a targeting chimera since it is highly complex and appears to be high risk. As a result we started thinking about a targeting chimera which could be synthesized by the cell itself. We contacted Prof. Dr. Claudine Kraft and set up a first meeting.
