itFISH: Enhanced Staining by Iterative Fluorescent In Situ Hybridization.
Fluorescent in situ hybridization (FISH) is an important tool for zebrafish research, particularly when observing the expression of two different genes in the same embryo. Peroxidase-catalyzed deposition of tyramide-conjugated dyes is a widely used and cost-effective approach to performing FISH. A major limitation of the technique is that it does not work well for weakly expressed genes. Here we present a method adapted from planarian research for use in zebrafish that provides a dramatic enhancement of weak staining. By iterating the antibody staining and development steps, a strong signal can be obtained from probes that were previously too weak to detect.