Subzonal insemination of a single mouse spermatozoon with a personal computer‐controlled micromanipulation system

Abstract

A personal computer-controlled micromanipulation system was developed for automatic injection of spermatozoa into the perivitelline space of mouse ova. A pair of three-dimensional hydraulic micromanipulators driven by pulse motors was used for this automatic system. The pulse signals that regulate the motors are initiated by the computer program, and these signals cause the micromanipulator to move the microtool precisely. The computer program was designed to perform the most effective movements of the sperm injection needle used during manual micromanipulation. Prior to the manipulation, the computer locates the tip of the injection needle and the end of the egg-holding pipette in the microscope field using image processing. The trajectory of the injection needle is determined according to these initial positions. Using this robotic system, subzonal insemination with a single mouse spermatozoon was attempted in a total of 143 ova. The sperm insertion was successfully completed in all cases without damaging any of the ova. Spermatozoa treated with ionophore A23187 and those without the treatment were used. The fertilization rate (68.8%) of the ova inseminated with treated sperm was significantly higher than that (37.5%) obtained with the nontreated sperm (P less than 0.05). These findings suggest the feasibility and potential for further applications of a robotic microinsemination system and, in addition, that a higher fertility rate in the subzonal insemination of mouse ova can be achieved with the ionophore treatment of spermatozoa.

A Fully Automated Robotic System for Microinjection of Zebrafish Embryos

“…Manual injection is not only slow; the laborious task of manual injection easily causes fatigue in injection technicians and hinders performance consistency and success rates. Efforts in automating cell injection have been continuous, resulting in a visually servoed system [7], a semi-automated system [8], and many tele-operated systems [9]–[13], to name just a few. These systems are limited in throughput and reproducibility as operator input (e.g., locating features and destinations) or operator involvement (e.g., switching from one cell to another or injector alignment) is still required.…”

A haptic training environment for the heart myoblast cell injection procedure

“…Traditional cell injection manipulation requires long training and the success rate is low for even an experienced scientist. However, with the aid of a computer controlled 3 DOF micro-robotic system, the successful rate is 80%, as demonstrated in Kobayashi et al [11]. Wang et al [12] also utilised a 3 DOF semi-automated micro-robotic system that combined with computer vision for sequential cell injection.…”

Pretreating porcine sperm with lipase enhances developmental competence of embryos produced by intracytoplasmic sperm injection

“…Since the first report of mammalian ICSI in rabbits (Hosoi et al, 1988;Iritani, 1988), transfer of embryos produced by ICSI has given rise to live cattle (Goto et al, 1990), mice (Kobayashi et al, 1992), sheep (Catt & Rhodes, 1995), horses (Squires et al, 1996), cats (Pope et al, 1998), and humans (Palermo et al, 1992). It has been proved that directly injecting a single spermatozoon into an oocyte can produce apparently normal offspring even though it by-passes a series of biological processes necessary for normal fertilization, such as the acrosome reaction, sperm capacitation, and membrane fusion.…”

“…ICSI operations can produce normal offspring regardless of spermatozoa concentration, morphology, and motility, as long as the sperm nucleus has an intact genetic identity. Since the first report of mammalian ICSI in rabbits (Hosoi et al , 1988; Iritani, 1988), transfer of embryos produced by ICSI has given rise to live cattle (Goto et al , 1990), mice (Kobayashi et al , 1992), sheep (Catt & Rhodes, 1995), horses (Squires et al , 1996), cats (Pope et al , 1998), and humans (Palermo et al , 1992). However, successful production of piglets using in vivo matured oocytes after ICSI was not reported until 2000 by Martin (2000).…”