In order to circumvent ethical, technical, and economic drawbacks regarding the use of animal serum in cell culturing, it is possible to adapt mammalian cells to serum‐free media. Nowadays, there are several serum‐free formulations available, including fully animal derived–free and chemically defined media, and different adaptation techniques. This article focuses on the gradual adaptation of a mammalian suspension cell culture to a chemically defined medium. The first step is to transfer the cells cultured in medium supplemented with fetal bovine serum (FBS) to a chemically defined medium of your choice, containing the same amount of FBS. The next steps consist of progressively reducing the amount of FBS, while monitoring cell growth and viability up to the complete elimination of FBS. This protocol has been successfully used to adapt THP‐1 cells to a chemically defined medium with similar maximum specific growth rate as those cultured with FBS. © 2019 by John Wiley & Sons, Inc. Basic Protocol: Gradual adaptation to chemically defined medium Alternate Protocol: Direct adaptation to chemically defined medium Support Protocol 1: Determining maximum specific growth rate of a cell culture Support Protocol 2: Cell freezing and thawing

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In vitro methods that can replace animal testing in the identification of skin sensitisers are now a reality. However, as cell culture and related techniques usually rely on animal-derived products, these methods may be failing to address the complete replacement of animals in safety assessment. The objective of this study was to identify the animal-derived products that are used as part of in vitro methods for skin sensitisation testing. Thus, a systematic review of 156 articles featuring 83 different in vitro methods was carried out and, from this review, the use of several animal-derived products from different species was identified, with the use of fetal bovine serum being cited in most of the methods (78%). The use of sera from other animals, monoclonal antibodies and animal proteins were also variously mentioned. While non-animal alternatives are available and methods free of animal-derived products are emerging, most of the current methods reported used at least one animal-derived product, which raises ethical and technical concerns. Therefore, to deliver technically and ethically better in vitro methods for the safety assessment of chemicals, more effort should be made to replace products of animal origin in existing methods and to avoid their use in the development of new method protocols.

Most rabies vaccines are based on inactivated virus, which production process demands a high level of biosafety structures. In the past decades, recombinant rabies virus glycoprotein (RVGP) produced in several expression systems has been extensively studied to be used as an alternative vaccine. The immunogenic characteristics

Insect cells are used for the expression of complex proteins in products such as vaccines and biopharmaceuticals. Physiology of a Drosophila melanogaster (lineage S2), transfected to stably express rabies virus glycoprotein (RVGP), was studied in batch culture and in a chemostat with serum-free medium. In batch mode, the system reached 3 × 10(7) cells mL(-1) with specific growth rate of 1.5 d(-1) with RVGP at 2.50 µg L(-1). When operated continuously, three well-defined steady states were achieved at dilution rates (D) of 0.8, 0.5 and 0.2 d(-1). The residual glucose and glutamine concentrations and the cell yields on glucose and glutamine decreased at lower D. High values of substrate consumption for maintenance may explain this variation in yields. The results indicated that glucose is not the limiting substrate of this process.