Related Experiment Video
Updated: Dec 31, 2025
Visualization of Macrophage Lytic Cell Death During Mycobacterial Infection in Zebrafish Embryos via Intravital Microscopy
Published on: January 9, 2019
Visualization of Macrophage Lytic Cell Death During Mycobacterial Infection in Zebrafish Embryos via Intravital
Liangfei Niu1, Cong Wang1, Kaile Zhang2
1Shanghai Public Health Clinical Center, Fudan University.
Abstract:
Zebrafish is an excellent model organism for studying innate immune cell behavior due to its transparent nature and reliance solely on its innate immune system during early development. The Zebrafish Mycobacterium marinum (M. marinum) infection model has been well-established in studying host immune response against mycobacterial infection. It has been suggested that different macrophage cell death types will lead to the diverse outcomes of mycobacterial infection. Here we describe a protocol using intravital microscopy to observe macrophage cell death in zebrafish embryos following M. marinum infection. Zebrafish transgenic lines that specifically label macrophages and neutrophils are infected via intramuscular microinjection of fluorescently labeled M. marinum in either the midbrain or the trunk. Infected zebrafish embryos are subsequently mounted on low melting agarose and observed by confocal microscopy in X-Y-Z-T dimensions. Because long-term live imaging requires using low laser power to avoid photobleaching and phototoxicity, a strongly expressing transgenic is highly recommended. This protocol facilitates the visualization of the dynamic processes in vivo, including immune cell migration, host pathogen interaction, and cell death.
Insights
This study details a zebrafish embryo protocol to visualize macrophage cell death during Mycobacterium marinum infection using intravital microscopy. This method aids in understanding host-pathogen dynamics and immune responses in vivo.
Area of Science:
- Immunology
- Developmental Biology
- Microbiology
Background:
- Zebrafish are ideal models for innate immunity research due to transparency and early-life reliance on innate immunity.
- The zebrafish Mycobacterium marinum (M. marinum) model is established for studying mycobacterial infections.
- Macrophage cell death is implicated in diverse outcomes of mycobacterial infections.
Purpose of the Study:
- To describe a protocol for observing macrophage cell death in zebrafish embryos infected with M. marinum using intravital microscopy.
- To visualize dynamic in vivo processes of host-pathogen interaction and immune cell behavior.
Main Methods:
- Infection of zebrafish embryos (transgenic lines labeling macrophages/neutrophils) via intramuscular microinjection of fluorescent M. marinum.
- Mounting infected embryos in low melting agarose for confocal microscopy.
- Utilizing X-Y-Z-T dimensions for dynamic imaging, recommending strongly expressing transgenics for low-laser power live imaging.
Main Results:
- The protocol enables visualization of macrophage cell death in real-time within infected zebrafish embryos.
- Dynamic immune cell migration, host-pathogen interactions, and cell death events are observable.
- Successful imaging of host immune responses to M. marinum infection.
Conclusions:
- This intravital microscopy protocol provides a powerful tool for studying innate immune cell death during mycobacterial infection in zebrafish.
- It facilitates a deeper understanding of the dynamic host-pathogen interactions and immune responses in vivo.
- The method is crucial for dissecting the role of macrophage cell death in infection outcomes.
