Standard specimens for stain calibration: application to Romanowsky-Giemsa staining.

J N Turner, B Weir, D N Collins
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引用次数: 5

Abstract

Standardized specimens with reproducible staining properties were fabricated from extracts of biological objects (bovine liver, nucleoprotamine and defatted muscle). The standard specimens were stained with two formulations of the Romanowsky-Giemsa stain (RG), using the same azure B and eosin Y. One formulation used methanol and Sorensen's buffer and the other DMSO and Hepes buffer as solvents. The standard specimens were stained either in the composite stain or in the individual dyes dissolved in the same solvents and at the same concentration as the composite stain. Solution spectroscopy demonstrated different spectra for the two formulations with some wavelength regions varying by more than an order of magnitude. The RG spectra were also very different from those of the individual dyes dissolved at the RG concentration in the respective solvents. The stained standard specimens were analyzed by microspectrophotometry and were found to have spectra similar to those of cell smears. Furthermore, the standard specimens were shown to be a repeatable substrate for stain uptake. The transmitted light intensity from random fields of the same standardized specimen varied +/- 5%. When specimens were stained at the same time, the specimen-to-specimen variation depended on preparation conditions and the measurement wavelength, but was as good as +/- 5% for some conditions. The quantitative stain performance of both formulations was studied and compared. The standardized specimens provide a tool for the quantitative study of staining processes and specimen preparation procedures and for stain calibration.

染色校正的标准标本:用于罗曼诺夫斯基-吉姆萨染色。
从生物对象(牛肝脏、核鱼精蛋白和脱脂肌肉)的提取物中制备具有重复性染色特性的标准化标本。标准标本用两种配方的罗曼诺夫斯基-吉姆萨染色剂(RG)染色,使用相同的天蓝色B和伊红y。一种配方使用甲醇和Sorensen缓冲液,另一种配方使用DMSO和Hepes缓冲液作为溶剂。标准标本在复合染色剂中染色,或在与复合染色剂相同的溶剂和浓度下溶解的单个染料中染色。溶液光谱学证明了两种配方的光谱不同,某些波长区域的变化超过一个数量级。RG光谱也与溶解在相应溶剂中RG浓度下的单个染料的光谱有很大的不同。用显微分光光度法分析染色后的标准标本,发现其光谱与细胞涂片相似。此外,标准标本被证明是一种可重复的基底,用于染色吸收。同一标准化试样随机场透射光强变化+/- 5%。当标本同时染色时,样品间的变化取决于制备条件和测量波长,但在某些条件下可达+/- 5%。研究并比较了两种配方的定量染色性能。标准化的标本为染色过程和标本制备程序的定量研究以及染色校准提供了工具。
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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