What is multiplex fluorescent mRNA in situ hybridization?
In situ hybridization is one of the techniques that form the bedrock of molecular developmental biology, with which patterns of gene expression in embryos can be visualized. The use of non-radioactive probes and hybridization to whole Drosophila embryos instead of tissue sections (Tautz and Pfeifle, 1989) radically simplified the technique while also achieving a much higher level of resolution. In this new method, probes labeled with the small molecule digoxigenin could be visualized after hybridization with immunochemistry, as shown in this diagram: After hybridization, probes are recognized by an anti-digoxigenin antibody conjugated with alkaline phosphatase. During incubation with the substrate NBT/BCIP, this enzyme deposits a brownish purple precipitate in the embryos, which then turns a deep blue color after exposure to ethanol. In this way, cells which have produced a particular mRNA are stained, and the pattern of stained cells across an entire embryo can be seen, such as the si
