Protocol for high-resolution 3D visualization of insect regenerating legs through micro-computed tomography

Appendage regeneration occurs within the opaque exoskeleton in arthropods, making it challenging to visualize the regenerative processes dynamically. In this protocol, we present a strategy to scan and capture the high-resolution details of microstructural tissues at certain regeneration points through micro-computed tomography (micro-CT). We describe steps for tissue preparation, fixation, critical point drying, micro-CT scanning, and 3D visualization. This approach promises significant utility in the field of regeneration, particularly in studies involving arthropods such as insects and crustaceans. For complete details on the use and execution of this protocol, please refer to Ren et al.¹.