3D + Time Imaging and Image Reconstruction of Pectoral Fin During Zebrafish Embryogenesis

Morphogenesis is the fundamental developmental process during which the embryo body is formed. Proper shaping of different body parts depends on cellular divisions and rearrangements in the growing embryo. Understanding three-dimensional shaping of organs is one of the basic questions in developmental biology. Here, we consider the early stages of pectoral fin development in zebrafish, which serves as a model for limb development in vertebrates, to study emerging shapes during embryogenesis. Most studies on pectoral fin are concerned with late stages of fin development when the structure is morphologically distinct. However, little is known about the early stages of pectoral fin formation because of the experimental difficulties in establishing proper imaging conditions during these stages to allow long-term live observation. In this protocol, we address the challenges of pectoral fin imaging during the early stages of zebrafish embryogenesis and provide a strategy for three-dimensional shape analysis of the fin. The procedure outlined here is aimed at studying pectoral fin during the first 24 h of its formation corresponding to the time period between 24 and 48 h of zebrafish development. The same principles could also be applied when studying three-dimensional shape establishment of other embryonic structures. We first discuss the imaging procedure and then propose strategies of extracting quantitative information regarding fin shape and dimensions.