Immunofluorescent Quantification of MIF in Larval Zebrafish Tailfin Transection.

The use of zebrafish in research is becoming increasingly popular, requiring the development of analytical techniques to suit the organism. One of these techniques is immunofluorescence staining, which allows the visualization and quantification of specific proteins of interest. The immunofluorescence staining technique involves the binding of primary and secondary antibodies (with attached fluorophores) to a protein of interest-in this case, macrophage migration inhibitory factor (MIF). Due to the small size and transparency of zebrafish larvae, this can be performed using the whole body, so that no sectioning equipment is required. Making the technique even more adaptable and user-friendly, the high genetic homology observed between humans and zebrafish often allows the application of antibodies raised against human antigens in zebrafish imaging. For example, there is a 79% amino acid sequence homology between human and zebrafish MIF, with previous imaging demonstrating clear similarities in the expression patterns between the two species. This chapter will provide an in-depth description of the methods involved in immunofluorescence staining of MIF in zebrafish larvae, using tailfin transection as the experimental, pro-inflammatory insult.