In a study of the metal distribution in a heavy metal tolerant fungus by means of X-ray microanalysis, the intensities of copper X-rays in a semi-thin section specimen of the cell cultivated in high copper concentration medium were found to be different between various fixation methods: glutaraldehyde fixation, glutaraldehyde-osmium vapor fixation and glutaraldehyde-osmium solution fixation. The copper content in the cell samples and the used fixative after the fixations were determined by atomic absorption spectrometry. After the fixations, the cell samples had approximately the same copper content. A copper signal was always detected together with an osmium signal. It was suggested that Kα X-ray emission of copper might be increased by Lβ 1 X-ray and the atomic number effect of osmium.