Prolonged in vivo imaging of Xenopus laevis

Background: While live imaging of embryonic development over long periods of time is a well established method for embryos of the frog Xenopus laevis, once development has progressed to the swimming stages, continuous live imaging becomes more challenging because the tadpoles must be immobilized. Current imaging techniques for these advanced stages generally require bringing the tadpoles in and out of anesthesia for short imaging sessions at selected time points, severely limiting the resolution of the data. Results: Here we demonstrate that creating a constant flow of diluted tricaine methanesulfonate (MS‐222) over a tadpole greatly improves their survival under anesthesia. Based on this result, we describe a new method for imaging stage 48 to 65 X. laevis, by circulating the anesthetic using a peristaltic pump. This supports the animal during continuous live imaging sessions for at least 48 hr. The addition of a stable optical window allows for high quality imaging through the anesthetic solution. Conclusions: This automated imaging system provides for the first time a method for continuous observations of developmental and regenerative processes in advanced stages of Xenopus over 2 days. Developmental Dynamics 243:1011–1019, 2014. © 2014 Wiley Periodicals, Inc.