Institute of Microbiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
Eur J Pharm Biopharm. 2010 Nov;76(3):514-24. doi: 10.1016/j.ejpb.2010.07.008. Epub 2010 Jul 16.
There is a wide range of techniques utilizing fluorescence of doxorubicin (Dox) commonly used for analysis of intracellular accumulation and destiny of various drug delivery systems containing this anthracycline antibiotic. Unfortunately, results of these studies can be significantly influenced by doxorubicin degradation product, 7,8-dehydro-9,10-desacetyldoxorubicinone (D*) forming spontaneously in aqueous environment, whose fluorescence strongly interfere with that of doxorubicin. Here, we define two microscopy techniques enabling to distinguish and separate Dox and D* emission based either on its spectral properties or on fluorescence lifetime analysis. To analyze influx and nuclear accumulation of Dox (free or polymer-bound) by flow cytometry, we propose using an indirect method based on its DNA intercalation competition with Hoechst 33342 rather than a direct measurement of doxorubicin fluorescence inside the cells.
有一系列广泛应用的技术利用阿霉素（Dox）的荧光，这些技术通常用于分析含有这种蒽环类抗生素的各种药物输送系统的细胞内积累和归宿。不幸的是，这些研究的结果可能会受到阿霉素降解产物 7,8-去氢-9,10-去乙酰基阿霉素酮（D*）的显著影响，D在水环境中会自发形成，其荧光会强烈干扰阿霉素的荧光。在这里，我们定义了两种显微镜技术，可以根据其光谱特性或荧光寿命分析来区分和分离 Dox 和 D的发射。为了通过流式细胞术分析游离或聚合物结合的 Dox 的内流和核积累，我们提出使用一种间接方法，该方法基于其与 Hoechst 33342 的 DNA 插入竞争，而不是直接测量细胞内阿霉素的荧光。
