Delgado C, Anderson R J, Francis G E, Fisher D
Molecular Cell Pathology Laboratory, Royal Free Hospital School of Medicine, University of London, United Kingdom.
Anal Biochem. 1991 Feb 1;192(2):322-8. doi: 10.1016/0003-2697(91)90543-3.
The partitioning of cells in aqueous two-phase systems formed by poly(ethylene glycol) (PEG) and dextran can be changed by incubating the cells with a PEG-modified antibody directed specifically against its surface. We have developed a new approach for immunoaffinity cell partitioning (IACP) in which the antibodies are first reacted with tresylated monomethoxy PEG (TMPEG) in sodium phosphate buffer, pH 7.5, the excess TMPEG is quenched by reaction with bovine serum albumin, and the resulting preparation is used directly for incubation with the cells without any isolation of the monomethoxyPEG (MPEG)-antibody conjugates. We have demonstrated the specificity of this IACP method by showing that MPEG-modified anti-human red blood cell antibody increases the partition of human erythrocytes from the interface to the PEG-rich top phase (up to 100%) but not the partitioning of either neutrophils or HL60 cells. Irrelevant antibodies do not affect the partitioning of red blood cells. The partitioning behaviors of erythrocytes and HL60 cells in mixtures varying from 75 to 10% red blood cells subjected to IACP are similar to those of the pure cell population, i.e., erythrocytes ca. 100% and HL60 cells 3% in top phase. Thus, the population of erythrocytes can be almost completely extracted into the top phase in a single step. The contaminant cells represent only a small percentage (less than 5% in most of the cases) of the cell mixture recovered in top phase. Both cell populations can be completely separated by countercurrent distribution (CCD).(ABSTRACT TRUNCATED AT 250 WORDS)
聚乙二醇（PEG）和葡聚糖形成的双水相系统中细胞的分配情况，可通过将细胞与特异性针对其表面的PEG修饰抗体孵育来改变。我们开发了一种免疫亲和细胞分配（IACP）的新方法，其中抗体首先在pH 7.5的磷酸钠缓冲液中与三羟甲基氨基甲烷磺酸化单甲氧基PEG（TMPEG）反应，过量的TMPEG通过与牛血清白蛋白反应淬灭，所得制剂可直接用于与细胞孵育，无需分离单甲氧基PEG（MPEG）-抗体缀合物。我们通过证明MPEG修饰的抗人红细胞抗体增加了人红细胞从界面到富含PEG的上相的分配（高达100%），但不影响中性粒细胞或HL60细胞的分配，从而证明了这种IACP方法的特异性。无关抗体不影响红细胞的分配。在75%至10%红细胞混合物中进行IACP处理时，红细胞和HL60细胞的分配行为与纯细胞群体相似，即上相中红细胞约为100%，HL60细胞约为3%。因此，红细胞群体可以在一步中几乎完全提取到上相中。在上相中回收的细胞混合物中，污染细胞仅占小百分比（大多数情况下小于5%）。两种细胞群体可通过逆流分配（CCD）完全分离。（摘要截短于250字）
