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.

中文翻译：

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线虫蠕虫秀丽隐杆线虫的自动机器人显微注射

线虫蠕虫 秀丽隐杆线虫是一种模式生物，广泛用于遗传学，发育，神经科学和衰老的生物学研究。显微注射是一种有效且被广泛采用的方法，可用于创建转基因蠕虫，对某些基因进行核糖核酸（RNA）干扰，并将不同类型的分子引入蠕虫体内的特定位置。基于微流控技术和机器人微操纵技术，我们开发了一种用于自动显微注射的机器人系统。秀丽隐杆线虫与传统的手动显微注射相比，注射速度大大提高，成功率更高。具有计算机控制的气动阀的双层微流控设备被开发出来，用于单蠕虫的自动片上装载，固定，注射和下游分选。提出了一种新的基于自动聚焦的接触检测算法，以沿着显微镜视场的深度方向找到最佳注射位置。针尖的方向和位置可以使用图像处理算法可靠地识别。通过对240个蠕虫进行实验，该系统演示了以6个蠕虫/分钟（9.97 s /蠕虫）的速度进行自动注射，预分选成功率达78.8％（分选后成功率：100％），是23倍以上速度比速度快1.6倍（0。熟练的操作员分别达到25蠕虫/分钟和30％的成功率。凭借卓越的性能，该系统将使新的大规模基因和分子筛选研究成为可能。秀丽隐杆线虫 传统的显微注射技术无法实现。 执业者须知 —在蠕虫生物学界，全世界有成千上万的研究实验室例行地应对蠕虫显微注射实验。本文旨在介绍我们用于高速蠕虫注入的自动化机器人系统的功能和性能。使用机器人系统，大量秀丽隐杆线虫可以将其装载到微流控设备中，以进行连续的蠕虫固定和注射。开发了用户友好的图形用户界面（GUI），以使操作员可以在计算机屏幕上监视注射过程，选择蠕虫体内的注射位置（通过计算机鼠标单击）以及引导系统（通过键盘输入）用于成功分选的蠕虫的下游分选，以进行进一步培养。鉴于其独特的功能，例如高注射速度，高自动化水平和高成功/成活率，该系统具有将蠕虫研究人员从繁琐的手动注射过程中解放出来的巨大潜力，并提供无与伦比的注射产量和一致性。