A relatively undemanding 3D reconstruction protocol using in situ hybridization on serial sections successfully visualized gene expression patterns in embryonic mouse hearts.
Describes an accessible 3D reconstruction protocol for visualizing gene expression patterns in embryonic development using in situ hybridization on serial sections.
The study of the genetic regulation of embryonic development requires the three-dimensional (3D) mapping of gene expression at the microscopic level. Despite the recent burst in the number of methods focusing on 3D reconstruction of embryonic specimens, an adequate and accessible 3D reconstruction protocol for the visualization of patterns of gene expression is lacking. In this communication we describe a protocol that was developed for the 3D visualization of patterns of gene expression determined by in situ hybridization (ISH) on serial sections. The method still requires tissue sectioning, due to penetration limits of the specific staining agents into whole embryo preparations. With regard to expenditure of resources, i.e., hardware, software, and time, the protocol is relatively undemanding. Because the variation between specimens requires the visualization of multiple specimens per stage, it was decided to "do more, less well." The current protocol, therefore, results in reconstructions of sufficient, but not the highest, quality. The use of the protocol is demonstrated on a series of serially sectioned mouse hearts, ranging from embryonic day 8.5 to 14.5. The myocardium of the hearts was identified by ISH using a mixture of specific mRNA probes and reconstructed.
Soufan et al. (Tue,) conducted a other in Cardiac development. 3D reconstruction protocol was evaluated on 3D visualization of patterns of gene expression. A relatively undemanding 3D reconstruction protocol using in situ hybridization on serial sections successfully visualized gene expression patterns in embryonic mouse hearts.