Journal of Microscopy (Oxford) 2001-01-01

A simple contrast enhancement by potassium permanganate oxidation for Lowicryl K4M ultrathin sections prepared by high pressure freezing/freeze substitution.

A Sawaguchi, S Ide, Y Goto, J Kawano, T Oinuma, T Suganuma

Index: J. Microsc. 201(Pt 1) , 77-83, (2001)

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Abstract

A simple contrast enhancement method is presented for Lowicryl K4M ultrathin sections prepared by high pressure freezing/freeze substitution. The sections were treated with an acidified potassium permanganate oxidizing solution followed by uranyl acetate and lead citrate staining. The method, designated KMnO4-UA/Pb staining, provided a much greater contrast in electron microscopy than conventional UA/Pb staining. In detail, the visibility of plasma membrane was especially improved and the nuclear heterochromatin, mitochondria and cytoplasmic ribosomes showed an adequate increase in electron density. In the mucous cells of rat Brunner's glands, the Golgi cisternae were well defined with the KMnO4-UA/Pb staining. Interestingly, the membranes of the intermediate compartments were moderately reactive to the KMnO4-UA/Pb staining, whereas the cis and the trans compartments were only faintly stained. It should be emphasized that the KMnO4 oxidation following colloidal gold labelling did not cause a remarkable reduction of immunogold labelling and the enhanced contrast helped us to examine the gold particles with high accuracy. This contrast enhancement method is highly promising, with the potential to become a useful tool for histochemical investigation, including immunocytochemistry with the Lowicryl K4M ultrathin sections prepared by high pressure freezing/freeze substitution techniques.