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目的建立一套有效可靠的检测方法,以测定阴离子脂质体介导的腺病毒和化疗药卡莫司汀共转运载体系统中腺病毒和卡莫司汀的含量。方法以钙离子融合法制备共转运载体复合物;根据腺病毒(Ad5)的hexon基因序列设计用于Taqman探针荧光定量聚合酶链式反应(PCR)的引物和探针序列,优化反应体系,从而准确测定腺病毒含量;优化色谱条件,建立针对共转运载体复合物检测卡莫司汀含量的高效液相色谱法。结果以荧光定量聚合酶链式反应法测得共转运载体复合物中腺病毒载药量为(23.2±1.8)%;高效液相色谱法测得卡莫司汀在共转运载体复合物中的载药量为(55±2.8)%。结论本实验成功建立了分别用于测定共转运载体系统中腺病毒和卡莫司汀含量的荧光定量聚合酶链式反应和高效液相色谱法;这两种检测方法操作简便、准确可靠,具有一定的普遍适用性。
OBJECTIVE: To establish an effective and reliable method for the determination of adenovirus and carmustine in the carmustine cotransporter system mediated by anionic liposomes. Methods The cotransporter vector was prepared by calcium ionophoresis. According to the sequence of hexon gene of adenovirus (Ad5), primers and probe sequences for Taqman probe were designed, and the reaction system was optimized. Thus accurately determining the content of adenovirus; optimizing the chromatographic conditions, and establishing a high performance liquid chromatography for detecting the content of carmustine for the cotransporter carrier complex. Results The amount of adenovirus loaded in the cotransporter complex was (23.2 ± 1.8)% as measured by fluorescence quantitative polymerase chain reaction. Carmustine was detected in the cotransporter complex by high performance liquid chromatography Drug loading was (55 ± 2.8)%. Conclusion In this study, we successfully established fluorescent quantitative polymerase chain reaction and high performance liquid chromatography respectively for the determination of adenovirus and carmustine levels in the cotransporter system. The two detection methods were simple, accurate and reliable, with Certain general applicability.