PREPARATION AND CHEMICAL CHARACTERISTICS OF HEMOGLOBIN-FREE GHOSTS OF HUMAN ERYTHROCYTES
PREPARATION AND CHEMICAL CHARACTERISTICS OF HEMOGLOBIN-FREE GHOSTS OF HUMAN ERYTHROCYTES
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DOI：
10.1016/0003-9861(63)90042-0
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发表时间：
1963-01-01
影响因子：
3.9
通讯作者：
MITCHELL, C
中科院分区：
文献类型：
--
作者：
DODGE, JT;HANAHAN, DJ;MITCHELL, C
在磷酸盐缓冲液中研究了溶血溶液的离子强度和 pH 值对人红细胞血影血红蛋白含量的影响，发现对血影中的血红蛋白结合有显着影响。缓冲液浓度在 10 到 20 理想毫渗摩尔 (imOsm) 之间、pH 值 5.8-8.0 时，可以最大程度地从血影中去除血红蛋白。在 20 imOsm 缓冲液中，血红蛋白与血影结合的最佳 pH 值在 5.8 到 5.9 之间。这些变量的影响表明血红蛋白与膜成分存在电物理相互作用。这项研究为比较现有的血影制备方法提供了基础，并为预测制备含有任何所需量的血红蛋白的血影所需的条件提供了一种方法。发现了可以通过一步溶血和洗涤制备无血红蛋白血影的条件。在 pH 7.4 的 20 imOsm 磷酸盐缓冲液中制备无血红蛋白的血影。鬼魂中基本上恢复了所有脂质，但非血红蛋白含氮物质丢失了。用于血红蛋白测定的吡啶血色原方法适用于通过使用 Soret 带 (418 mu) 进行吸光度测量来测量极少量的血红蛋白。
The effects of the ionic strength and pH of the hemolyzing solution on the hemoglobin content of human erythrocyte ghosts were studied in phosphate buffers and found to have a pronounced influence upon hemoglobin binding in the ghosts. Buffer concentrations between 10 and 20 ideal milliosmolar (imOsm), at pH values 5.8-8.0, resulted in maximum hemoglobin removal from ghosts. The pH optimum for hemoglobin binding to ghosts was between 5.8 and 5.9 in a 20 imOsm buffer. The influence of these variables suggest an electrophysical interaction of hemoglobin with membrane constituents. This study provides a basis for comparison of existing methods for ghost preparation, as well as a means for prediction of the conditions required for preparation of ghosts containing any desired amount of hemoglobin. Conditions were found that allowed the preparation of hemoglobin-free ghosts by single-stage hemolysis and washing. Hemoglobin-free ghosts were prepared in 20 imOsm phosphate buffer at pH 7.4. Essentially all the lipid was recovered in the ghosts, but non-hemoglobin nitrogen-containing substances were lost. The pyridine hemochromogen method for hemoglobin determination was adapted for the measurement of very small quantities of hemoglobin through use of the Soret band (418 mu) for absorbancy measurements.