Related Experiment Video
Updated: Jul 15, 2026
Mouse Round Spermatid Injection
Published on: January 26, 2024
Piezo-actuated mouse intracytoplasmic sperm injection (ICSI)
Naoko Yoshida1, Anthony C F Perry
1Laboratory of Mammalian Molecular Embryology, RIKEN Center for Developmental Biology, 2-2-3 Minatojima Minamimachi, Chuo-ku, Kobe 650-0047, Japan.
Abstract:
The mouse is a genetically tractable model organism widely used to study mammalian development and disease. However, mouse metaphase II (mII) oocytes are exquisitely sensitive and intracytoplasmic sperm injection (ICSI) with conventional pipettes generally kills them. This problem can be solved with piezo-actuated micromanipulation, in which the piezo-electric effect (crystal deformation in response to an externally applied voltage) propels a microinjection needle tip forward in a precise and rapid movement. Piezo-actuated micromanipulation enhances the penetration of membranes and matrices, and mouse ICSI is a major application. Here we describe a comprehensive, step-by-step mouse piezo ICSI protocol for non-specialists that can be completed in 2-4 h. The protocol is a basic prelude to multiple applications, including nuclear transfer cloning, spermatid injection, blastocyst injection, mII transgenesis, and streamlining micromanipulation in primates and livestock. Moreover, piezo ICSI can be used to obtain offspring from 'dead' (non-motile) sperm, enabling trivial sperm freezing protocols for mouse strain storage and shipment.
Insights
A new protocol for mouse intracytoplasmic sperm injection (ICSI) using piezo-actuated micromanipulation significantly improves oocyte survival rates. This method enables efficient genetic manipulation and offspring production, even from non-motile sperm.
Area of Science:
- Reproductive Biology
- Developmental Biology
- Genetics
Background:
- Mouse oocytes are sensitive to conventional intracytoplasmic sperm injection (ICSI) methods, often resulting in cell death.
- Developing robust micromanipulation techniques is crucial for advancing mammalian reproductive technologies and genetic studies.
- Existing ICSI protocols pose significant challenges for mouse oocyte viability and subsequent applications.
Purpose of the Study:
- To present a detailed, step-by-step protocol for mouse piezo-actuated ICSI suitable for non-specialists.
- To demonstrate the utility of piezo ICSI for various reproductive and genetic applications in mice.
- To establish a simplified method for mouse strain preservation using cryopreserved non-motile sperm.
Main Methods:
- Utilized piezo-actuated micromanipulation for precise and rapid microinjection needle movement.
- Developed a comprehensive 2-4 hour protocol for mouse intracytoplasmic sperm injection (ICSI).
- Applied the protocol to achieve successful oocyte penetration and subsequent embryo development.
Main Results:
- Piezo-actuated micromanipulation significantly enhances membrane and matrix penetration during ICSI.
- The protocol facilitates high survival rates for mouse metaphase II oocytes post-ICSI.
- Successful offspring production was achieved using non-motile sperm, simplifying cryopreservation.
Conclusions:
- Mouse piezo ICSI offers a highly effective and accessible method for assisted reproduction in laboratory mice.
- This technique serves as a foundation for advanced applications like nuclear transfer cloning and transgenesis.
- The protocol streamlines micromanipulation for broader applications in mammalian and potentially non-mammalian species.
