摘要
目的 :为深入研究骨髓巨噬细胞的生物学功能 ,探索一种简便的骨髓巨噬细胞的分离 ,纯化和培养的方法。方法 :分离获取大鼠骨髓细胞 ,在 6 0 %DMEM培养液 ,2 0 %马血清 ,2 0 % (v/v)L92 9培养上清的诱导条件下进行体外贴壁培养 ,6～ 7天时获得纯度较高的贴壁细胞。采用倒置显微镜下观察生活状态、Wright′s染色光镜观察、电子显微镜观察检测贴壁细胞形态学 ;酸性磷酸酶及非特异性酯酶染色测定细胞内酶的表达 ;吞噬鸡红细胞及墨汁颗粒实验检测其吞噬功能 ;免疫细胞化学染色观察细胞表面标志等生物学技术鉴定培养的贴壁细胞性质。结果 :获得高纯度的巨噬细胞 ,且具有良好的吞噬功能 ,并且具备巨噬细胞的形态特征 ,特有的水解酶类及特有的表面标志 -CD68。结论 :本法是一种简易实用的体外分离、纯化、培养骨髓巨噬细胞的方法。
Objective:To study the biological functions of bone marrow macrophage (BMM Φ) and to establish the methodology of separation, purification, cultivation and identification of BMM Φ.Methods:Using the techniques of anchorage-dependent culture of separated rat bone marrow cell (rBMC) in DMEM culture media (contain 20% horse serum,20% (v/v)L 929 conditioned media) in vitro,a lot of purified anchor cells were obtained, and these cells were identified with specifically biological marker of macrophage, such as 1, morphologicalobservation:invert phase contrast microscopy, light and electron microscopy;2, enzyme cytochemistry:acid phosphatase (ACP),α-acetic acid naphthol esterase (α-ANE);3,phagocytic experiment:phagocytosis of chicken erythrocytes and prepared Chinese ink;4,immunocytochemistry: surface specific antigen of macrophage ( CD 68 stain).Results:The cells were purified having functional satisfactory macrophage according to morphological observation,enzyme cytochemistry, phagocytic experiment and immunocytochemistry.Conclusion:This is a simple and easy method for separation, purification, cultivation and identification of rat marrow macrophage.
出处
《重庆医科大学学报》
CAS
CSCD
2003年第4期436-439,共4页
Journal of Chongqing Medical University
关键词
骨髓巨噬细胞
分离
纯化
鉴定
Bone marrow macrophage
Separation
Purification
Identification
