Malpighian Tubule Dissection from an Adult Fly: Isolating an Osmoregulatory and Excretory Organ

- Begin with an anesthetized adult fly in a dissecting dish. Hold the fly by the thorax with forceps and open the posterior end of the abdomen using a second pair of forceps. Since adult flies have two pairs of malpighian tubules or MTs that converge onto the gut through common ureters, tug the hind gut out of the fly to release the MTs. The posterior pair of MTs will be freed first as they encircle behind gut. Continue tugging the gut to free the anterior MTs and then pinch them off at the ureter to separate them from the fly.

To mount the tubules, slide a glass rod under the ureter and transfer the MTs to poly-L-lysine- coated slide. Affix the dissected organs by pressing the ureter down and then carefully sweep the rod over the MTs. The poly-L-lysine on the slide will promote attachment by increasing the number of positively charged sites available for the negatively charged cells of the tubules to bind. In the following protocol, we will dissect MTs from an adult female fly and mount them for ex vivo live imaging in a perfusion system.

- To begin this procedure, position two sets of soldering helping hands clamps on the imaging microscope stage with one clamp on each side. Next