Universal robot for automated microinjection with applications in transgenesis and cryopreservation

Abstract

Microinjection is the process of injecting a small amount of solution into biological organisms at a microscopic level using a glass micropipette. It is a widely utilized technique with a wide range of applications in both fundamental research and clinical settings. However, microinjection is an extremely laborious and manual procedure, which makes it a critical bottleneck in the field and thus ripe for automation. In this thesis, we introduce a simple computer vision-guided robot that uses off-the-shelf components to fully automate the microinjection procedure in different model organisms. The robot uses machine learning models that have been trained to detect individual embryos on agar plates and serially performs microinjection at a particular site in each detected embryo with no human interaction. We deployed three such robots operated by expert and novice users to perform automated microinjections in zebrafish (Danio rerio) and Drosophila melanogaster. We conducted survivability studies to better understand the impact of microinjection on zebrafish embryos and the fundamental mechanisms by which microinjection affects zebrafish embryos. We were able to use the robot to examine the speed of the micropipette, the volume of the microinjectant, the micropipette geometry, and the rate of the volume delivered. These results helped us in determining the optimum settings for automated microinjection into zebrafish embryos. We used transgenesis studies to compare microinjection efficiency to manual microinjection utilizing optimum settings for automated microinjection. Further, we demonstrated that robotic microinjection of cryoprotective agents in zebrafish embryos significantly improves vitrification rates and post-thaw survivability of cryopreserved embryos compared to manual microinjection, opening the door to large-scale cryo-banking of various aquatic species on an industrial scale. We anticipate that this robotic microinjection can be readily adapted to other organisms and applications.