Hybridization to tissues in suspension for whole-mount FISH in Drosophila

Hybridization to some Drosophila tissues, such as embryos or egg chambers, is easily performed in suspension. The reaction is set up in 0.5-mL polymerase chain reaction (PCR) tubes and the solutions are changed by allowing the tissue to settle to the bottom of the tube, aspirating the liquid carefully, and replacing it with 400-500 µL of a new solution. This is by far the most convenient way to perform the hybridization and requires a minimum volume for all washing steps. The denaturation step is also easy to perform consistently because the sample can be heated and annealed in a thermal cycler or conventional water bath. This protocol describes hybridization to tissues in suspension for whole-mount fluorescent in situ hybridization (FISH) in Drosophila.