Using the “Drop Method” as Part of a Refined Euthanasia Protocol: Welfare and Practical Considerations

Inhalant anesthesia, typically with isoflurane, is recommended for mice prior to euthanasia. While vaporizers are the standard delivery method, they may not be available in all facilities. An alternative is the “drop method”, in which liquid isoflurane is dispensed directly into an induction chamber. This method is practical, cost-effective, and well-suited for terminal procedures where precise anesthetic control is not required. Here, we outline our application of the drop method for mouse isoflurane anesthesia, including the induction procedure, and considerations including induction chamber design, isoflurane dosage calculations, and the effects of varying concentrations on mouse behavior and welfare. We also address animal welfare and human safety concerns, providing strategies to minimize occupational isoflurane exposure. We conclude that with appropriate chamber design, accurate dosing, and careful monitoring, the drop method serves as an effective and humane alternative for anesthesia prior to euthanasia, helping to refine euthanasia procedures in facilities lacking vaporizer access.