The influence of mode of fixation, type of fixative and vehicles on the same rat liver: a morphometric/stereologic study by light and electron microscopy.

The influence on cellular and subcellular structure was studied by comparing perfusion versus immersion fixation with cacodylate buffered glutaraldehyde, osmium or glutaraldehyde immersion fixation with two vehicles, phosphate and cacodylate buffer. The study was performed in the same animal using the left liver lobe for the four immersion fixation experiments and the remaining lobes for perfusion. The preparative steps were done under identical conditions in one operation. The mode of fixation was the most influential factor with clear (15%) increases for the volume of hepatocytes (mainly their cytoplasm), and the mitochondria (30%), when comparing immersion to perfusion fixation. This mitochondrial enlargement was particularly displayed in the organelles' average profile area, being more than doubled (211%) in immersion fixation. Similarly high profile enlargements (150%) were also found after osmium immersion fixation. Changes in nuclei were minor compared to the cytoplasm. There was a good correlation between the volume changes of the hepatocyte cytoplasm and the mitochondria (r = 0.85, p less than 0.01). Qualitative results revealed that there exists a zone of good electron microscopic preservation 2-3 cell layers beneath the block edge in immersion fixed material, whereas perfusion fixed tissue displays a homogeneous fine structure.