Em localization of atpase on microtubules of isolated cilia from Tetrahymena vorax.

J T Fitzsimons, G A Kerkut

Index: Comp. Biochem. Physiol. A. Comp. Physiol. 74(3) , 739-43, (1983)

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Abstract

1. Isolated cilia were prepared from Tetrahymena vorax using the local anaesthetic dibucaine in the deciliation step. 2. ATPase was cytochemically localized on microtubules of isolated cilia using the Washstein-Meisel incubation; deposition of lead phosphate indicated the sites of enzyme activity. 3. Mild fixation conditions gave optimum localizations. Satisfactory results were attained using 0.5% glutaraldehyde with a fixation time of 30 min. 4. An increase in ATPase activity, as judged by lead phosphate precipitation, was observed when cytochemical incubations were increased from 5 min to 1 hr. An incubation time of 15 min gave optimum results. 5. No advantage was gained with incubation times over 1 hr as diffusion of reaction product may occur. 6. No ATPase activity was observed in control incubations where the enzyme substrate ATP was omitted. 7. Purified cilia preparations provide useful starting material for the study of microtubular ATPase.