Ref ID : 4080
Eugene J. Hoffman, Douglas A. Price, and James A. Krumpelman; Isolation of Axopodial Axonemes from Mass-cultured Echinosphaerium nucleofilum. J.Protozool. 30(2):371-375, 1983
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The heliozoan Echinosphaerium nucleofilum produced about 75x10E3 floating cells per 19-cm culture dish per day when fed the green flagellate Chlorogonium elongatum. This method yields enough cells for usable quantities of subcellular fractions. Heliozoa were lysed in a detergent mixture containing stabilizing reagents, and axonemal bundles of axopodial microtubules were isolated from the lysate by differential centrifugation. Polyacrylamide gel electrophoresis in sodium dodecylsulfate showed two prominent bands tentatively designated alpha- and beta-tubulin. Apparent molecular weights were 51.8x10E3 and 48.1x10E3, respectively. As assayed by electron microscopy of negatively stained whole mounts, the microtubules bundles splintered readily, although glycerol tended to inhibit this fraying. Intermicrotubule bridges could be observed in some axonemal splinters.