Automated Robotic Microinjection of the Nematode Worm Caenorhabditis elegans

The nematode worm Caenorhabditis elegans is a model organism widely used in biological research on genetics, development, neuroscience, and aging. Microinjection is an effective and widely adopted method to create transgenetic worms, perform ribonucleic acid (RNA) interference of certain genes, and introduce different types of molecules into specific locations inside a worm body. Based on microfluidics and robotic micromanipulation techniques, we develop a robotic system for automated microinjection of C. elegans with greatly improved injection speed and success rate over traditional manual microinjection. A double-layer microfluidic device with computer-controlled pneumatic valves is developed for automated on-chip loading, immobilization, injection, and downstream sorting of single worms. A new autofocusing-based contact detection algorithm is proposed to find the optimal injection position along the depth direction of the microscope field of view. The direction and location of the needle tip are reliably identified using an image processing algorithm. Through experiments on 240 worms, the system demonstrates automated injection at a speed of 6 worms/min (9.97 s/worm) with a presorting operation success rate of 78.8% (postsorting operation success rate: 100%), which are more than 23 times faster and 1.6 times higher than the speed (0.25 worm/min) and success rate (30%) of a proficient human operator, respectively. With the superior performance, this system will enable new large-scale gene-and molecule-screening studies on C. elegans that cannot be fulfilled by the conventional microinjection technique. Note to Practitioners-In the worm biology community, there are thousands of research laboratories worldwide that routinely cope with worm microinjection experiments. This article aims to present the functionality and performance of our automated robotic system for high-speed worm injection. Using the robotic system, a large number of C. elegans can be loaded into the microfluidic device for continuous worm immobilization and injection. A user-friendly graphical user interface (GUI) is developed to allow an operator to monitor the injection process on a computer screen, select the injection location inside the worm body (through computer mouse clicking), and direct the system (through keyboard input) for downstream sorting of the successfully injected worms for further culture. Given its unique features, such as high injection speed, high level of automation, and high success/survival rates, this system holds great potential to liberate worm researchers from the tedious manual injection process and provide unparalleled injection throughput and consistency.