Commentary – Non-animal-derived antibodies: addressing ethical and scientific needs

Abstract

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In two recently published articles, we reported the production, isolation and characterization of new non-animal-derived antibodies targeting the clinically significant human CD33 and human PD-1 proteins (Dhungana et al., 2026a, 2026b). We evaluated the ability of these antibodies to recognize their targets across a range of experimental applications, including comparisons with commercially available antibodies, showing that the performance of these new antibodies is at least comparable to animal-derived antibodies. These findings augment the body of evidence that non-animal-derived antibodies can address longstanding challenges in biomedical research and support a move away from animal-derived antibodies (Modi, 2025; Barroso et al., 2020) in alignment and compliance with the EU Directive on the Protection of Animals Used for Scientific Purposes (European Parliament and Council of the European Union, 2010). To understand the context for this work and set the stage for steps that can accelerate this process, it is important to first define the terminology surrounding antibodies from different sources for clarity, consistency, and for their potential impact on animal replacement. These are as follows:

• Recombinant antibodies are produced in vitro using recombinant DNA technology, rather than animal-derived hybridomas, using defined genetic sequences to express the antibody protein in cell lines. While the production of the antibody is animal-free (not taking into account the possible use of animal-derived biomaterials in cell culture), the discovery phase may or may not involve animals: antibody sequences may be derived from in vitro screens, such as phage display, but may also be derived from antibodies that have been produced via the immunisation of animals.
• Immunisation-dependent recombinant antibodies are antibodies produced recombinantly, but discovered and initially isolated via animal immunisation and associated animal procedures described below.
• Non-animal-derived antibodies have been both discovered and produced using animal-free methods with no animal immunisation involved. However, their production may involve animal-derived biomaterials such as fetal bovine/calf serum (FBS/FCS).

Incorporated in the laws and institutional guidelines governing animal use in research in many countries are key ethical principles first described by Russell and Burch (Russell and Burch, 1959). The “Three Rs” include replacement, reduction, and refinement. Adherence to these principles is legally mandated in much of Europe by EU Directive 2010/63/EU (European Parliament and Council of the European Union, 2010). The principle of replacement has received particular priority due to the animal welfare issues involved in antibody production, as well as the explosive growth of recombinant technology.

Species commonly used in antibody production include rabbits, mice, rats, goats, sheep, horses and chickens, among others. Production processes vary, but typically animals undergo immunization with a target antigen, along with adjuvants and sometimes booster immunizations to potentiate immune responses. Small bleeds follow to ensure the animals are producing antibodies sufficiently, followed by larger bleeds to harvest greater amounts of antibodies. Animals may be bled multiple times, and/or may be killed and their spleens removed to create cell cultures (hybridomas) that are used to produce commercial antibodies. An animal may be used for several years.

Ethical concerns are accompanied by scientific issues. Publications over the past decade have reviewed the limitations of animal-derived antibodies and the advantages of non-animal-derived antibodies. Groff et al. suggested that animal-derived antibodies are a major contributor to the reproducibility crisis in science and outlined efforts by industry and regulators to promote the transition to non-animal-derived antibodies through policy and funding changes (Groff et al., 2024). An editorial in Nature highlighted case studies suggesting that up to two-thirds of the approximately 8 million animal-derived antibodies on the market fail to perform adequately, including instances in which they do not bind their target molecule(s) (Kwon, 2024). In 2020, the EURL ECVAM Scientific Advisory Committee (ESAC) published a comprehensive report, concluding that non-animal-derived antibodies are scientifically valid and often superior to animal-derived antibodies (with improved purity, activity, specificity, affinity, stability, and reproducibility), recommending that animal antibodies not be used or authorized unless there is a “robust and legitimate justification” (Barroso et al., 2020).

Since their initial development in the mid-1970s, non-animal-derived antibodies have offered the possibility of replacing animal-derived antibodies. In theory, the procedures are straightforward. Genetic sequences for specific antibodies are identified and optimized, then inserted into expression vectors, which enable the production of the antibodies when these recombinant vectors are introduced into host cell cultures. The antibodies are then purified and tested for specificity and affinity.

The impact on animal replacement differs based on how and when recombinant technologies are used, for example: if animal immunisation was used in discovery or not; if immunisations remain part of the methodology, even if these are reduced; if polyclonal or ascites production is replaced; if animal-derived biomaterials are utilised and to what degree. In other words, “recombinant” and “non-animal-derived” do not always mean “animal-free”. Ultimately, whatever the details, recombinant techniques aim to facilitate the production of more consistent and reproducible antibodies and easier downstream modification, which have significant impacts on reproducibility and on the waste of scientific materials, time, funding, and animals’ lives. The use of sequence-defined antibodies, therefore, whatever their status and derivation, is a positive step in many respects.Technological progress in recent years is such that replacement of animal use is no longer a theoretical prospect; rather, the development of a wide range of high-quality non-animal-derived antibodies has become a reality. Non-animal-derived antibodies currently represent roughly one quarter of the 100 most-cited antibodies in recent years (Cell Signaling Technology, 2026). Nonetheless, the adoption of recombinant antibodies has not reached its full potential, reflecting barriers related to awareness, availability, and perceived cost (Groff et al., 2024). To encourage greater use of non-animal-derived antibodies, experts have developed practical recommendations, including the following:

• Government agencies, research funders, publishers, industry, and academic institutions should prioritize the replacement of animal-derived antibodies and establish clear guidance and mechanisms to monitor implementation and compliance (Bradbury & Plückthun, 2015).

• Scientific organizations, academic institutions, and training programs should develop educational materials to increase awareness that non-animal-derived antibodies are widely available and that their use can improve research quality, reproducibility, and animal welfare.
• Governments, research funders, and industry partners should expand public-private partnerships and increase investment in the development, validation, and dissemination of non-animal-derived antibodies to improve accessibility.
• Researchers, manufacturers, publishers, and repositories should ensure that antibodies are identifiable (Barroso et al., 2020), preferably by sequence, through the use of ABCD and Research Resource Identifiers (RRIDs) (Lima et al., 2020; Kwon, 2024), and encourage their validation to be published in peer-reviewed journals.
• Researchers should make greater use of antibody databases, including CiteAb, ABCD and the Recombinant Antibodies & Mimetics Database, to identify validated recombinant reagents for specific applications (Kahn et al., 2024; Modi, 2025).

Non-animal-derived antibodies have proven value. Increased awareness, investment, and prioritization of these tools can accelerate their incorporation in scientific research.