免疫磁珠法分选小鼠肝脏Kupffer细胞及其特性观察

Isolation and purification of mouse liver Kupffer cells by immunomagnetic bead separation

目的:建立免疫磁珠法,分离纯化小鼠肝脏Kupffer细胞并进行功能鉴定。方法:采用原位肝脏酶灌注、制备肝非实质细胞悬液、加入PE标记的大鼠抗小鼠F4/80一抗,然后结合山羊抗大鼠IgG Dynabeads免疫磁珠分选Kupffer细胞,流式细胞仪检测其纯度并检测其吞噬功能。结果:在经过胶原酶Ⅳ型消化、免疫磁分选,获得小鼠肝Kupffer细胞产量约为(8.05±0.34)×106/鼠肝,细胞纯度为(96.6±0.7)%,细胞活力大于90%。结论:应用免疫磁珠法能有效地分离纯化小鼠肝脏Kupffer细胞,为体外进一步研究提供了高纯度和活力的Kupffer细胞群。

Objective ：To establish the way of irnmunomagnetic bead separation for isolating and purifying mouse liver Kupffer cells and exploring their phagocytic function. Methods：The techniques included the processes of liver perfusion in situ, preparation of non-parenchymal cells suspension, isolation using anti-mouse F4/80 antibody and sheep anti-rat IgG Dynabeads. Finally, flow cytometry was used to detect the purity and phagocytic function. Results： By collagenase Ⅳ digestion and immunomagnetic bead separation, Kupffer cells were obtained in the amount of （8.05±0. 34） × 10^6/mouse liver. The purity was （96.6 ± 0.7 ） % and more than 90% of cells were viable. Conclusion： Mouse Kupffer cells can be isolated and purified by immunomagnetic bead separation, which makes it possible to further the study of this cell population in vitro.