Benchaib M, Delorme R, Pluvinage M, Bryon P A, Souchier C
Analytical Cytology Laboratory, Claude Bernard University, Lyon, France.
Histochem Cell Biol. 1996 Aug;106(2):253-6. doi: 10.1007/BF02484409.
Fluorescein isothiocyanate (FITC) is largely used in immunofluorescence methods. We propose to analyse the quality of some recent fluorochromes using image analysis. Fluorochromes tested include FITC and dichlorotriazinylaminofluorescein (DTAF), dipyrrometheneboron difluoride (BODIPY), Rhodol Green and cyanine 2. RAMOS cells were immunolabelled against the proliferating cell nuclear antigen (PCNA) revealed by the biotin-streptavidin technique. Slides were mounted in anhydrous glycerol or in buffered glycerol (pH 7.0 or pH 8.5). No antifading medium was added. Cell fluorescence emission intensity and bleaching characteristics were measured. Rhodol Green exhibited the highest fluorescence intensity and the best photobleaching resistance. Although BODIPY also resisted well during the photobleaching assay, its fluorescence intensity was weak. FITC, DTAF and cyanine 2 showed intermediate fluorescence intensity and a fast decay of fluorescence. Among the green emitting fluorochromes tested, Rhodol Green appeared to be the best.
异硫氰酸荧光素（FITC）在免疫荧光方法中被大量使用。我们建议使用图像分析来分析一些近期荧光染料的质量。所测试的荧光染料包括FITC、二氯三嗪基氨基荧光素（DTAF）、二氟硼二吡咯（BODIPY）、罗丹明绿和花青2。通过生物素-链霉亲和素技术对RAMOS细胞进行抗增殖细胞核抗原（PCNA）的免疫标记。载玻片用无水甘油或缓冲甘油（pH 7.0或pH 8.5）封固。未添加抗褪色介质。测量细胞荧光发射强度和漂白特性。罗丹明绿表现出最高的荧光强度和最佳的抗光漂白性。尽管BODIPY在光漂白试验中也表现出良好的抗性，但其荧光强度较弱。FITC、DTAF和花青2表现出中等荧光强度和快速的荧光衰减。在所测试的发射绿色荧光的荧光染料中，罗丹明绿似乎是最好的。
