大鼠肺泡巨噬细胞原代培养技术的改良措施

目的改良原代肺泡巨噬细胞（alveolar macrophage,AM）培养技术,提高AM培养效果。方法SD大鼠,在体支气管灌洗肺获取AM,通过预先注射空气、14号注射器针头回收灌洗液、低速离心、种板前多次吹打等改良措施分离和培养AM。倒置显微镜连续观察AM的形态变化,台盼蓝拒染实验鉴定AM存活率,瑞氏-姬姆萨染色鉴定AM纯度。结果AM体外培养后40 min即见细胞贴壁;2～3天时,细胞形态多样,呈圆形、不规则形、梭形等形态,胞体变大,胞质丰富。台盼蓝拒染实验示AM存活率≥95%,瑞氏-姬姆萨染色示AM纯度≥95%。结论本实验改良了原代AM培养的技术方法,提高了原代AM培养效果。

Modifications of the primary culture technique for rat alveolar macrophages

Objective To modify the techniques for primary culture of highly purified the primary rat alveolar macrophages（AMs）.Methods AMs were isolated in vivo and cultured with such modified techniques as pre-injection of air and reclamation of brochoalveolar lavage fluid with 14-bugle syringe needle,low-speed centrifugation and multi-blowing before plating on 6-well culture plates.The morphological changes were observed under the inverted microscope.The cell survival rate was detected with trypan blue dye exclusion test,and cell purity was determined by Wright-Giemsa staining.Results In vitro,AMs adhered to plastic surface within 40 min;2-3 days later,cell morphology presented mixed shapes,i.e.,circular,irregular and spindle shapes.The trypan blue test showed a survival rate ≥95% of the AMs and Wright-Giemsa staining indicated a purity ≥95% of the AMs.Conclusion The modified techniques employed in our experiment had better results for the isolation and primary culture of AMs.