Removal of insect basal laminae using elastase.
We have used the enzyme elastase to remove the basal lamina of epithelia from two insects: the upper Malpighian tubules of Rhodnius prolixus and imaginal discs of Drosophila melanogaster. Removal of the basal lamina was confirmed using scanning and transmission electron microscopy. Use of the technique on the Malphighian tubules of Rhodnius reveals for the first time the three-dimensional organization of the circumferential folds of the basal plasma membrane. Elastase is much more effective in removing the basal lamina than are the enzymes hyaluronidase, collagenase, and chymotrypsin, either alone or in combination. Following elastase treatment, cells of the Malpighian tubules dissociate with only mild mechanical agitation into single, viable cells. Treatment with elastase removes the basal laminae of imaginal discs of Drosophila and accelerates evagination as has been previously described for trypsin. To obtain single cell preparations from elastase-treated imaginal discs, mechanical stirring in Ringer low in Ca2+ was required. In addition to its usefulness in cell isolation, elastase treatment allows examination of the effect of removal of basal laminae on the physiology and development of insect epithelia.
D000818:Animals; D001485:Basement Membrane / Q000187:drug effects / Q000648:ultrastructure*; D002469:Cell Separation / Q000379:methods*; D004331:Drosophila melanogaster / Q000648:ultrastructure; D007313:Insecta / Q000166:cytology*; D008317:Malpighian Tubules / Q000648:ultrastructure; D008854:Microscopy, Electron; D010196:Pancreatic Elastase / Q000494:pharmacology*; D012239:Rhodnius / Q000648:ultrastructure; D014921:Wings, Animal / Q000648:ultrastructure
D010196:Pancreatic Elastase