Individual microtubules viewed by immunofluorescence and electron microscopy in the same PtK2 cell

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Abstract
PtK2 cells [rat kangaroo] were grown on Au, grids and treated with Triton X-100 in a microtubule stabilizing buffer. The resulting cytoskeletons were fixed with glutaraldehyde and subjected to the indirect immunofluorescence procedure using monospecific tubulin antibodies. Grids were examined first by fluorescence microscopy, and the display of fluorescent cytoplasmic microtubules was recorded. The grids were then stained with uranyl acetate and the display of fibrous structures recorded by EM. The display of cytoplasmic microtubular structures in the light microscope and the EM can be compared within the same cytoskeleton. The results show a direct correspondence of the fluorescent fibers in the light microscope with uninterrupted fibers of diameter .apprx. 550 .ANG. in the EM. This is the diameter reported for a single microtubule decorated around its circumference by 2 layers of antibody molecules. Under optimal conditions immunofluorescence microscopy can visualize indivudal microtubules.