The use of a line scan ratemeter for the X-ray microanalytic evaluation of membrane-bound histochemical endproducts.

Scanning electron microscopy Pub Date : 1986-01-01
I Nagy, V Nagy
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Abstract

Although X-ray microanalysis represents a useful tool for identifying electron dense histochemical end-products, quantitative microanalytic measurements are seriously hampered in the case of the activities of certain membrane-bound enzymes. For example, the electron histochemical methods revealing K+-dependent pNPPase activity result in a very fine, granular reaction product of lead phosphate. Therefore microanalytic, densitometric of similar evaluations of the reaction, even in semiquantitative terms are not practical by the usual procedures. This paper describes a method of X-ray microanalysis of thick sections (0.5 micron) processed for K+-pNPPase, where a sufficient amount of lead is present for X-ray microanalytic determination. The analysis is performed in the line scan mode on transversely cut membrane profiles by means of the line scan ratemeter of an EDAX System F. This yields quantitative data on the relative lead concentrations in the vicinity of the cell membrane. A method is proposed for calculation of relative enzyme activities based on the Pb-signal of the ratemeter curve and the average "noise"-level of the cytoplasm, containing also non-specifically bound lead. This method avoids the necessity of measuring the section thickness; it may be useful for a variety of purposes in the electron microscopic histochemistry of membrane-bound enzymes.

使用线扫描速率计对膜结合的组织化学终产物进行x射线微分析评价。
尽管x射线微分析是识别电子致密组织化学最终产物的有用工具,但在某些膜结合酶的活性情况下,定量微分析测量严重受阻。例如,电子组织化学方法揭示K+依赖的pNPPase活性导致非常细的颗粒状磷酸铅反应产物。因此,微分析、密度学对反应的类似评价,即使是半定量的,用通常的方法也是不实际的。本文描述了一种用于K+-pNPPase加工的厚切片(0.5微米)的x射线微分析方法,其中足够量的铅存在于x射线微分析测定中。通过EDAX系统f的线扫描速率计,以线扫描模式对横向切割的膜剖面进行分析,从而产生细胞膜附近相对铅浓度的定量数据。本文提出了一种计算相对酶活性的方法,该方法基于速率计曲线上的铅信号和含有非特异性结合铅的细胞质的平均“噪声”水平。这种方法避免了测量截面厚度的必要性;它可能在膜结合酶的电镜组织化学中有多种用途。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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