MS on EVs from the YSL of 3dpf zebrafish embryos

We set up a zebrafish model to study EVs in vivo by light microscopy, using CD63-pHluorin as a marker. To verify if exosomes from zebrafish have a comparable composition as human exosomes, and to validate the use of CD63 as a marker, we first analysed the compositon of EVs isolated from in vitro cultured fibroblasts (AB.9 ATCC), and could detect the presence of conventional exosome markers, including (zebrafish) CD63. In live embryos, we could track a population of exosomes from their source (the yolk syncytial layer (YSL)) to their final destination by live microscopy. To assess the composition of these exosomes, we dissociated YSL:CD63-pHluorin expressing 3dpf embryos and isolated exosomes from the supernatant and enriched for EVs from the YSL. This was compared to a background of total EVs isolated the control fish (non-expressing).