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

The zebrafish brain consists of neurons - nerve cells that play a prominent role in processing information. Additionally, it comprises a population of supporting immune cells, including macrophages and microglia, that inhabit the interneuronal space.

To isolate all these types of brain cells from a zebrafish, first, take anesthetized zebrafish larvae in a Petri dish containing appropriate media.

Visualize the zebrafish under a stereomicroscope and transect the larval heads above the yolk sac to keep the brain intact.

Transfer the excised heads into a homogenizer filled with pre-chilled media, placed on ice. Low temperature helps prevent degradation of brain cells.

With a pestle, homogenize the zebrafish brain tissue. Homogenization disintegrates the extracellular matrix and initiates the dissociation of neurons, macrophages, and microglia from the brain tissue.

This step also separates myelin - a lipid-rich insulating layer - from neurons.

Pass the homogenate through a strainer to entrap large cell clumps.

Centrifuge the filtrate to pelletize the cells and myelin sheath fragments. Discard the debris-containing supernatant.

Resuspend the pellet in an appropriate density gradient med