Brain Cell Isolation from Zebrafish Larvae: A Technique to Obtain Intact Neurons, Macrophages, and Microglia from Zebrafish Brain Tissue

Abstract

Source: Mazzolini, J. et al. Isolation and RNA Extraction of Neurons, Macrophages and Microglia from Larval Zebrafish Brains. J. Vis. Exp. (2018)

This video describes a protocol to isolate neurons, macrophages, and microglia from larval zebrafish brains by mechanically disrupting the brain tissue. The purified cell populations can further be used for genomic analysis.

Protocol

All procedures involving human participants have been performed in compliance with the institutional, national and international guidelines for human welfare and have been reviewed by the local institutional review board. 1. Homogenization NOTE: All steps are performed 4 °C. Add 1.5 mL Tricaine (15 mM) to 90 mm Petri dishes containing 50 larvae in 50 mL of E3 embryo medium treated with 200 µM PTU to termin…

Materials

1-phenyl 2-thiourea (PTU)	Sigma	P7629
Hepes	Gibco	15630-056
D-Glucose	Sigma	G8644-100ML
HBSS 1X	Gibco	14170-088
Percoll	GE Healthcare	17-0891-02
HBSS 10X	Gibco	14180-046
DPBS 1X	Gibco	14190-094
Tricaine (MS222)	Sigma	A5040
Sterilin standard 90 mm petri dishes	ThermoFisher	101VIRR
Surgical micro-scissors	Fine Science Tools	15000-00
3 mL Pasteur plastic bulk pipette	SLS	PIP4206
Glass homogenizer	Wheaton	357538
Sterilin standard 55 mm petri dishes	ThermoFisher	P55V
Percoll	GE Healthcare	17-0891-02
40 µm cell strainer	Falcon	352340
50 ml polypropylen conical tube	Falcon	352070
Centrifuge	Eppendorf	5804 R
Normal goat serum (NGS)	Cell Signalling	5425S
35 μm cell strainer cap	BD	352235