小鼠肺泡巨噬细胞的分离及鉴定  

Isolation and identification of alveolar macrophages in mice

作　　者：黄燕霞[1] 许娜[1,2] 赵红[2] 孙成彪 刘洋[1] 陈华鑫[1] 万家余[1] 刘文森[1] 金宁一[1] 

机构地区：[1]军事医学科学院军事兽医研究所,吉林长春130122 [2]吉林医药学院,吉林吉林132000

出　　处：《中国生物制品学杂志》2016年第4期413-416,共4页Chinese Journal of Biologicals

基　　金：国家自然科学基金资助项目(31372487)

摘　　要：目的分离小鼠肺泡巨噬细胞(alveolar macrophage,AM),并鉴定其纯度和吞噬活性。方法采用肺泡灌洗法分离小鼠AM,分别通过瑞氏-姬姆萨染色法和Diff-Quik染色法鉴定AM的纯度;将FITC标记的大肠埃希菌与AM按不同比例混合(10∶1、15∶1、20∶1)后孵育不同时间(20、25、30 min),荧光倒置显微镜下观察AM的吞噬活性。结果两种染色方法染色时,阳性细胞百分率均在94%以上,Diff-Quik染色法细胞阳性率略高于瑞氏-姬姆萨染色法,染色时间较瑞氏-姬姆萨染色法短,且背景干净无杂质。小鼠AM的吞噬率和吞噬指数随着FITC标记的细菌与AM比例的增加而增大,且随孵育时间的延长呈先升高再降低的趋势。当细菌与细胞比例为20∶1,孵育时间为25 min时,吞噬率和吞噬指数达最大,分别为(63.67±4.04)%和1.41±0.07。结论贴壁培养法可获得较高纯度的小鼠AM,Diff-Quik染色法鉴定巨噬细胞纯度优于瑞氏-姬姆萨染色法,其可代替瑞氏-姬姆萨染色法,成为一种快速、高效的鉴定巨噬细胞纯度的方法。Objective To isolate alveolar macrophages（AMs）of mice and determine their purity and phagocytic activity.Methods The AMs of mice were isolated by alveolar lavage, and determined for purity by Wright-Giemsa staining and Diff-Quik staining. FTIC-labeled E. coli was mixed with AMs at various ratios（10 ∶ 1, 15 ∶ 1 and 20 ∶ 1） and incubated for 20, 25 and 30 min separately, then observed for phagocytic activity by inverted fluorescent microscopy. Results Both the positive rates of AMs stained by two methods were more than 94%, while that by Diff-Quik staining was slightly lower than that by Wright-Giemsa staining. However, the time for Diff-Quik staining was shorter than that for Wright-Giemsa staining, while the background was clean without impurities. Both the phagocytic rate and phagocytic index of AMs increased with the increasing ratio of FTIC-labeled E. coli to AMs, and showed a tendency of increase at first then decrease with the increasing time for incubation, while reached the peak values of（63. 67 ± 4. 04）% and 1. 41 ± 0. 07 respectively when the ratio was 20 ∶ 1 and the incubation time was 25 min. Conclusion Highly purified mouse AMs were obtained by adherent culture. Diff-Quik staining was superior to Wright-Giemsa staining in determination of purity of macrophages, which might be used as a rapid and highly effective method for determination of purity of macrophages instead of Wright-Giemsa staining.

关 键 词：小鼠肺泡巨噬细胞 Diff-Quik染色法 吞噬活性 FITC标记大肠埃希菌 

分 类 号：Q-34[生物学]