149suspension to solubilize the agarose and cool in a 37 °C water
bath before use.- Dissecting stereomicroscope with transmitted illumination,
providing at least 50× magnification with adjustable zoom
objective. - A pair of Dumont #5 watchmaker forceps with “standard” tip
(see Note 3). - 3 mL graduated Pasteur pipettes for embryo transfer (see
Note 4 ). - Mineral oil for scaling injection drop size.
- Calibration slide with 0.05 or 0.1 mm intervals.
- Concentrated bleach.
- Standard microscope slides.
- 1 mm diameter capillary tubes without filaments, 10 cm long.
- Micropipette puller (see Note 5).
- Pneumatic microinjector with micromanipulator.
- Fluorescence microscope with 4× air, 10× air, and 60× oil
objectives. - Glass bottom dish for inverted microscopy (see Note 6).
- 100 mm petri dishes and 96-well plates with lids.
- Suitable centrifuge tubes for bacterial suspension handling and
centrifugation (1.5 and 50 mL).
3 Methods
Figure 1 shows a timeline of embryonic zebrafish infection experi-
mental procedure involving adult fish mating, collecting eggs, pos-
sible MO injections, culturing of bacterial strains, injections into
zebrafish blood circulation, and following the experiment until
5 days post-fertilization (dpf).Fig. 1 Timeline of embryonic zebrafish infection experimental procedure. The timeline represents the typical
zebrafish infection experiment, starting from setting up adult fish for eggs until terminating the experiment at
5 dpf (legal protection age in the UK when grown at 28 °C)
Infection Model in Zebrafish