一种用于红细胞膜蛋白的等电聚焦方法及其在二维电泳中的应用。

An isoelectric focusing procedure for erythrocyte membrane proteins and its use for two-dimensional electrophoresis.

作者信息

Dockham P A, Steinfeld R C, Stryker C J, Jones S W, Vidaver G A

出版信息

Anal Biochem. 1986 Feb 15;153(1):102-15. doi: 10.1016/0003-2697(86)90067-9.

Abstract

Procedures are described and evaluated for one-dimensional isoelectric focusing of erythrocyte membrane dissolved in lysine, urea, and Triton X-100 without using sodium dodecyl sulfate (SDS) and for two-dimensional electrophoresis with SDS in the second dimension. The membrane was completely dissolved, most of the proteins including the anion porter(s) entered the focusing gel, and complex, well-resolved patterns were seen. Ampholines, 2-mercaptoethanol, or SDS in the applied sample each seriously reduced focusing resolution and phenylmethylsulfonyl fluoride blurred the patterns. The two-dimensional patterns showed more and sharper spots than did patterns obtained from membrane initially dissolved with SDS. Anion porter spots were seen with both procedures. However, major cytoskeletal proteins were much less well recovered with the former procedure than with the latter.

摘要

本文描述并评估了在不使用十二烷基硫酸钠（SDS）的情况下，对溶解于赖氨酸、尿素和 Triton X - 100 中的红细胞膜进行一维等电聚焦的方法，以及在第二维使用 SDS 进行二维电泳的方法。膜完全溶解，大多数蛋白质包括阴离子转运蛋白进入聚焦凝胶，并观察到复杂且分辨率良好的图谱。应用样品中的两性电解质、2 - 巯基乙醇或 SDS 均严重降低聚焦分辨率，苯甲基磺酰氟使图谱模糊。二维图谱比最初用 SDS 溶解膜所获得的图谱显示出更多且更清晰的斑点。两种方法均可见阴离子转运蛋白斑点。然而，与后一种方法相比，前一种方法回收的主要细胞骨架蛋白要少得多。