CAJ | 학술논문

用免疫黏附和差速离心法分离和纯化小鼠胎肺Ⅱ型细胞,所得细胞纯度达(90±2)%,产量为每5只小鼠胎肺收获(23.6±7)×10~6个细胞.原代培养的小鼠胎肺Ⅱ型细胞在12～24 h 开始贴壁,在36～48 h呈指数生长,72 h后生长减缓,4～5 d左右细胞开始变形分化.免疫荧光鉴定胎肺Ⅱ型细胞表面活性蛋白C(SP-C)染色阳性,透射电镜观察到细胞内板层小体,细胞中proSP-C蛋白在培养48 h后表达较高.体外病毒转染成功,可见强烈荧光表达.成功建立了小鼠胎肺Ⅱ型上皮细胞的培养模型.
용면역점부화차속리심법분리화순화소서태폐Ⅱ형세포,소득세포순도체(90±2)%,산량위매5지소서태폐수획(23.6±7)×10~6개세포.원대배양적소서태폐Ⅱ형세포재12～24 h 개시첩벽,재36～48 h정지수생장,72 h후생장감완,4～5 d좌우세포개시변형분화.면역형광감정태폐Ⅱ형세포표면활성단백C(SP-C)염색양성,투사전경관찰도세포내판층소체,세포중proSP-C단백재배양48 h후표체교고.체외병독전염성공,가견강렬형광표체.성공건립료소서태폐Ⅱ형상피세포적배양모형.
Differential centrifugation and specific immunosorption were used to isolate and purify fetal mouse alveolar epithelial type Ⅱ cells (AEC Ⅱ). The cells purity was (90±2)% and quantity was (23.6±7)×10~6 in five fetal mice lungs. AEC Ⅱ adhered after cultured for 12～24 h. Following 36～48 h in culture, cells appeared exponential growth. After 72 h,the growing rate decreased gradually. On days 4～5 in culture,cells began to differentiate and transmogrify. The expression of SP-C in AEC Ⅱ was positive by immunofluorescence and lamellarbodies were revealed under electron microscope. The quantity of proSP-C in culture cells was increased gradually and got to the high value when cultured for 48 hours. The adenovirus transfection to AEC Ⅱ cells was successful and EGFP expression could be detected. This study was successful to develop the primary culture technology and identification method for AEC Ⅱ.