MICROFLUORIMETRIC STUDY OF TRANSLATIONAL DIFFUSION IN ERYTHROCYTE-MEMBRANES
MICROFLUORIMETRIC STUDY OF TRANSLATIONAL DIFFUSION IN ERYTHROCYTE-MEMBRANES
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DOI：
10.1016/0005-2736(74)90085-6
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发表时间：
1974-01-01
期刊：
影响因子：
--
通讯作者：
BAHR, W
中科院分区：
文献类型：
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作者：
PETERS, R;PETERS, J;BAHR, W
描述了一种可以定量研究单细胞膜中平移扩散的方法。用异硫氰酸荧光素标记人红细胞，然后溶血，产生正常形状的鬼影和强荧光。应用十二烷基硫酸钠聚丙烯酰胺凝胶电泳发现，很大一部分异硫氰酸荧光素与红细胞膜蛋白结合。在荧光显微镜下，单个鬼被暴露在一束强烈的光线下，每次只有一半的鬼被漂白。通过显微测量，研究了荧光物质是否会从膜的未漂白部分扩散到漂白部分，反之亦然。然而，在室温下20分钟的测量时间内，无法检测到这种扩散的显著程度。为了定量地评价实验数据，对球面扩散方程进行了求解，并对得到的解在两个半球上进行了积分。通过这些方法，从实验数据中得出了3·10−12cm2/s的值，作为20°-23°C时异硫氰酸荧光素标记的化合物在红细胞膜上扩散系数的上限。
A method is described which permits quantitative study of translational diffusion in the membranes of single cells. Human erythrocytes were labelled with fluorescein isothiocyanate and then hemolyzed, which yielded ghosts of normal shape and strong fluorescence. By application of sodium dodecylsulfatepolyacrylamide gel electrophoresis it was found that a very large part of fluorescein isothiocyanate was bound to the proteins of the erythrocyte membrane. In a fluorescence microscope, single ghosts were exposed to a sharply bounded intensive beam of light in such a manner that in each case only one half of the ghost was bleached. By microscopic measurements it was studied whether fluorescent material would diffuse from the unbleached part of the membrane into the bleached part and vice versa. However, within the measuring time of 20 min at room temperature a significant degree of such a diffusion could not be detected. In order to evaluate the experimental data quantitatively, the diffusion equation for a spherical surface was solved, and the obtained solution furthermore was integrated over the hemispheres. By these means a value of 3 · 10−12cm2/s was derived from the experimental data as an upper limit for the diffusion coefficient of fluorescein isothiocyanate-labelled compounds in the erythrocyte membrane at 20°–23°C.