Histochemical and immunocytochemical evaluation of copper and metallothionein in the liver and kidney of copper‐loaded rats

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Abstract
Histochemical methods do not always show a good correlation with analytical measurement of copper content and consequently immunoreactive staining techniques for metaliothionein (MT) have recently been employed for the differential diagnosis of copper-associated diseases. This study compares histochemical with immunocytochemical methods for the assessment of copper status. Male rats were fed a high copper (1 g/kg) diet for 16 weeks and killed sequentially during this period. The livers and kidneys were analysed for copper and zinc (atomic absorption spectrophotometry), and sections were stained with rubeanic acid and rhodanine for copper and for immunoreactive MT using the DNP localization system. Immunoreactive stains for MT corresponded better with copper content than histochemical stains and were more sensitive, albeit less selective, indicators of copper accumulation. Moreover, major differences in intracellular staining were apparent between the two methods, attributed to differences in copper binding and microcompartmentalization of metal.