应用定量分析法与图像观察法对光敏剂细胞内分布的对比研究  

Contrast study on the subcellular localization of photosensitizers by quantitative analysis and image observation method

作　　者：戴维德[1] 曾晶[2] 李晓松[2] 刘凡光[2] 顾瑛[2] 

机构地区：[1]卫生部北京医院超声科,北京100730 [2]解放军总医院激光科

出　　处：《山西医科大学学报》2009年第11期989-991,F0003,共4页Journal of Shanxi Medical University

基　　金：国家自然科学基金资助项目(60078021)

摘　　要：目的探讨定量分析法在光敏剂亚细胞定位研究中的应用特点,并与图像观察法进行比较。方法传代培养A549肺癌细胞,分别与光敏剂血卟啉单甲醚孵共同孵育2,12,24h。选择四种特异性细胞器荧光探针Rhodamine-123、Luciferyellow、DiOC6(3)和BODIPY分别标记细胞内线粒体、溶酶体、内质网和高尔基体,倒置荧光显微镜和致冷CCD探测HMME和探针荧光图像。观察细胞中光敏剂和荧光探针各自的荧光图像并分析光敏剂在细胞中的分布特点。定量计算探针荧光高强度区域与细胞区域内的HMME荧光均量比IB/IA。结果图像观察法显示,孵育2,12,24h,细胞内HMME几乎不进入细胞核,而主要弥散分布于细胞质及各细胞器。定量结果显示:A549肺癌细胞内,2h时IB/IA由高到低为:高尔基体、溶酶体、线粒体、内质网;12h时IB/IA由高到低为:高尔基体、溶酶体、内质网、线粒体;24h时IB/IA由高到低为:高尔基体、内质网、溶酶体、线粒体;高尔基体组IB/IA随时间上升。结论应用定量分析法较图像观察法更可以准确反映出HMME在细胞内分布随着孵育时间变化的特点。Objective To compare and evaluate the application value of quantitative analysis and image observation method in determi- ning the subcellular localization of photosensitizers. Methods A549 lung cancer cells were incubated with HMME for 2,12 and 24 h. Four subcellular organelles were labeled by fluorescent probes, and the fluorescence images of HMME and probes were detected using the inverted fluorescence microscope and cooled CCD. IB/IA was calculated quantitatively, which represents the ratio of HMME fluorescence intensity in region with high probe fluorescence intensity and in the cell. Results The images showed that, at three incubation times, HMME in both kinds of cells had diffuse distribution in cytoplasm and organelles, with little nuclear stain. The quantitative results showed that arranged in descending order of IB/IA in A549 lung cancer cells at 2 h,they were Golgi, lysosome, mitochondria, and ER in turn. At 12 h ,IB/IA was detected from high to low in Golgi ,lysosome ,ER and mitochondria. At 24 h ,IB/IA decreased in Golgi ,ER,lysosome, and mitochondria in turn. The IB/IA in Golgi increased with incubation time. Conclusion The characteristics of intracellular distribution of HMME along with incubating time can be more accurately embodied by quantitative analysis than by image observation method.

关 键 词：光敏剂 亚细胞定位 图像 定量分析 

分 类 号：R734.2[医药卫生—肿瘤]