単一ネフロン各部位別の代謝特性等の微量定量法

概要

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Since a single nephron is a functional unit of the kidney, individual microdissec ted segments from the nephron would be ideal tissue samples for investigations on renal pharmacology. Several micromethods have enabled researchers to analyze the biochemical and pharmacological characteristics of these nephron segments. Miniaturized cuvettes containing microliter volumes of samples can be applied for general procedures of photometry like Lowrys protein determination. Fluorometry becomes a more sensitive method when enzymatic cycling systems of NAD/NADH or NADP/NADPH are combined, which have been used for assays of enzyme activities or substrate contents in minute biological samples having tissue proteins less than 1 μg. The microchemiluminescence procedure has been successfully utilized for cellular ATP content or oxygen radical generation. Radioimmunoassay can be used to determine endogeneous components such as cyclic nucleotides, eicosanoids, etc. Continuous gradient polyacrylamide microgels prepared in 5- to 10-μl capillaries have made it possible to quantify the intranephron distribution of cytochrome P-450, xanthine oxidase and superoxide dismutase. As an example of the modern techniques, microscopic fluorometry using Fura-2AM has been established to identify agonist-induced cytosolic free calcium transients.