Temperature Gradient Assay: A Method to Test Temperature Preference in Drosophila Larvae

- To test the temperature preference of Drosophila larvae, create a temperature gradient by heating one side of an aluminum assay plate and cooling the other. Aluminum has high thermal conductivity, meaning the heat is transferred efficiently, creating a stable linear temperature gradient. After the system has equilibrated, record the temperatures across the plate in same-sized temperature zones, then load the center of the plate with clean larvae.

Cover the entire system to eliminate interference from bright light and start the trial. Larvae distribute through the gradient, avoiding aversive zones that are too warm or too cold, and settle in zones of favorable temperatures. This behavior is called thermotaxis. To assess the larvae's temperature preference, record the number of animals in each temperature zone.

In the example protocol, we will see how to set up a single-direction temperature gradient between 18 and 28 degrees Celsius, and how larvae are handled during the assay.

- To prepare a single-directional gradient, first, create a humid ambient environment for the assay. Place two aluminum blocks connected to separate water baths on wet paper towels 10 centimeters apart. The