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Scientists are using in vitro methods to answer important research questions and implementing strategies to maximize the reliability and human relevance of these methods. One strategy is to replace the use of fetal bovine serum (FBS)-an undefined and variable mixture of biomolecules-in cell culture media with chemically defined or xeno-free medium. In this study, A549 cells, a human lung alveolar-like cell line commonly used in respiratory research, were transitioned from a culture medium containing FBS to media without FBS. A successful transition was determined based on analysis of cell morphology and functionality. Following transition to commercially available CnT-Prime Airway (CELLnTEC) or X-VIVO™ 10 (Lonza) medium, the cells were characterized by microscopic evaluation and calculation of doubling time. Their genotype, morphology, and functionality were assessed by monitoring the expression of gene markers for lung cell types, surfactant production, cytokine release, the presence of multilamellar bodies, and cell viability following sodium dodecyl sulphate exposure. Our results showed that A549 cells successfully transitioned to FBS-free media under submerged and air-liquid-interface conditions. Cells grown in X-VIVO™ 10 medium mimicked cellular characteristics of FBS-supplemented media while those grown in CnT-Prime Airway medium demonstrated characteristics possibly more reflective of normal human alveolar epithelial cells.

Many companies and global regulatory programs have expressed the intent to move away from in vivo animal testing to new approach methods (NAMs) as part of product safety assessments. NAMs, which include non-animal approaches for testing and assessment – from computer-based modeling to in chemico or in vitro models – allow faster data gener­ation with potentially greater relevance to humans while avoiding animal use. To monitor progress implementing NAMs, each organization first must define what is in scope, starting with the definition of “animal” (e.g., mammals, vertebrates) and applicable studies (e.g., animals used for “in-house” experiments, at contract research organizations, as part of envi­ronmental monitoring). Next, organizations must establish baseline animal use, including defined rules for inclusion/ exclusion of animals that ensure consistency in future assessments. Lastly, organizations must establish metrics for animal savings based on the utility of NAM data. This paper presents one approach to establish “animal use” metrics in a toxi­cology program at The Dow Chemical Company. The premise of our program is that most NAM information has value for animal savings, but the value depends on how data are used (e.g., research and development, screening, or regulatory requirements) and the level of certainty for internal decision-making. This manuscript provides metrics on the impact of NAMs, allowing a quantitative assessment of animal use numbers over time, accountability for resources spent on NAM development, and identification of areas where NAM development is still needed. This approach can be refined for use at other organizations.

Antibodies are used in a range of research, diagnostic, and regulatory applications. Traditional methods for producing such reagents involve the immunization of animals, which introduces variability into the methods that use them and is not aligned with efforts to replace and reduce animal use. Experts from academia, biotechnology, government, and animal protection organizations met December 3, 2019, at the National Institutes of Health in Bethesda, MD, USA to discuss the status of development and use of animal-free recombinant antibodies and their potential to replace antibodies derived from animals. This paper summarizes the discussion and the actions that resulted to facilitate increased production and use of animal-free recombinant antibodies.

The development and regulatory approval of ectoparasiticides, including flea and tick control products, involves decades-old methods and the use of large numbers of animals to evaluate toxicity and efficacy. Animals also are used to rear (breed and feed) fleas and ticks for later use in testing. Non-animal methods for regulatory-required testing and rearing currently exist and, with further development, others could soon become available. Here we provide an overview of the state-of-the-science of non-animal methods for rearing and regulatory-required efficacy testing of flea and tick control products. Several remaining challenges as well as recommendations on the steps needed to replace animals in the evaluation of these products are discussed.