Welfare-Oriented Manual Restraint Technique for Substance Administration and Blood Sampling in Mice.

Animal testing remains essential in biomedical research, contributing significantly to understanding disease mechanisms and evaluating the efficacy and safety of drugs and chemical substances, with rodents among the most frequently used laboratory animals. Among the most common interventions in laboratory practice are handling and immobilizing restraint during procedures such as injections and blood sampling, which are routinely performed on conscious mice without sedation or anesthesia. Proper restraint is necessary to ensure the safety of both the animal and the handler. However, traditional restraint methods, such as hand fixation or restrainer tubes, are known to induce acute stress responses in mice, thereby negatively affecting animal welfare and potentially influencing experimental outcomes. To promote refinement within the framework of the 3Rs principle and to improve animal welfare, this protocol describes the use of a newly developed application tunnel for injections and blood sampling in mice. The tunnel provides a less harsh yet still secure method of restraint compared with conventional techniques. This welfare-oriented restraint method can be easily implemented for standard procedures, such as intraperitoneal and intravenous injections and blood sampling, while minimizing stress and improving handling conditions for laboratory mice.