Isolation of Intestinal Macrophage Subpopulations for High-Quality Total RNA Purification in Zebrafish.

Methods Protoc

Fish Immunology Laboratory, Facultad Ciencias de la Vida, Universidad Andrés Bello, Avenida República 330, Santiago 8370186, Chile.

Published: May 2024

Intestinal macrophages have been poorly studied in fish, mainly due to the lack of specific molecular markers for their identification and isolation. To address this gap, using the zebrafish (:EGFP) transgenic line, we developed a fluorescence-activated cell sorting strategy (FACS) that allows us to isolate different intestinal macrophage subpopulations, based on GFP expression and morphological differences. Also, we achieved the purification of high-quality total RNA from each population to perform transcriptomic analysis. The complete strategy comprises three steps, including intestine dissection and tissue dissociation, the isolation of each intestinal macrophage population via FACS, and the extraction of total RNA. To be able to characterize molecularly different macrophage subpopulations and link them to their functional properties will allow us to unravel intestinal macrophage biology.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC11130952PMC
http://dx.doi.org/10.3390/mps7030043DOI Listing

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