Development of Separation Methods for Biological Macromolecules and Cells Using Nonsynchronous Coil Planet Centrifuge

1. 采用非同步螺旋行星离心机（CPC）研究了水-水聚合物相体系分离蛋白质的最佳分离条件和螺旋柱的行星运动对分离效率的影响。采用三种不同体积（11,24和39ml）的螺旋柱进行分离，两相体系由12.5% （w/w）聚乙二醇1000和12.5% （w/w）磷酸二氢钾组成，样品包括细胞色素C、肌红蛋白和溶菌酶。采用39 mL多层线圈，在800转/分和10转/分的从头到尾洗脱模式下，分离效果最好，其中偏心线圈在相同的实验条件下也得到了类似的结果。进一步的实验还表明，在转子转速为CCW的情况下，顺时针（CW）旋转下流动相线圈或逆时针（CCW）旋转上流动相线圈时，从头到尾的洗脱效果最好。非同步CPC法洗脱血细胞成分非同步CPC法应用于生理缓冲液分离血细胞成分。绵羊血液样品的最佳分离条件为连续旋转（CCW）和连续旋转（CW），采用头尾洗脱模式。在600-1000 rpm的高速旋转下，将羊和人的血液样品完全分离并洗脱成两个峰，其中在0 rpm的转速下洗脱除红细胞外的所有细胞成分，如血小板和白细胞，在10 rpm的线圈旋转下洗脱红细胞。羊血的正常洗脱模式为800转/分，人血样品为700转/分。

1. Protein separation using the nonsynchronous coil planet centrifuge (CPC)The nonsynchronous CPC was employed to investigate the optimal separating conditions and the effect of planetary motion of coiled column on the partition efficiency using the protein separation with aqueous-aqueous polymer phase systems. The separation was examined using three kinds coiled column with each different volume (11, 24 and 39 mL), a two-phase system composed of 12.5% (w/w) polyethylene glycol 1000 and 12.5% (w/w) dibasic potassium phosphate and a set of stable proteins for samples including cytochrome C, myoglobin and lysozyme. The best separation was obtained using the 39 mL multilayer coil at the head to tail elution mode with the revolution at 800 rpm and the rotation at 10 rpm, where the similar result was obtained in the eccentric coil under same experimental conditions. The further experiments also revealed that the best result was obtained in the head to tail elution mode by the clockwise (CW) coil rotation for the lower mobile phase or by the counterclockwise (CCW) coil rotation for the upper mobile phase, both under CCW revolution of the rotor.2. Elutriation of blood cell components by the nonsynchronous CPCThe nonsynchronous CPC was applied to the separation of blood cell components using a physiological buffer solution. The optimal condition for the separation of sheep blood samples was performed at the CCW revolution and the CW rotation of coiled column by the head to tail elution mode. Under the high speed of revolution of 600-1000 rpm, sheep and human blood samples were each completely separated and eluted into two peaks, where all cell components other than erythrocytes such as platelets and leukocytes were eluted at 0 rpm and erytbrocytes at 10 rpm of coil rotation. The normal elution pattern was achieved at 800 rpm of revolution for sheep blood and at 700 rpm for human blood samples.