Evaluation study on distribution of drugs in cells and tissues (immunocytochemistry)

各种定量化学分析方法主要用于药物的药理研究，主要利用动物的尿液、血液和组织匀浆。此外，通常用于药物药理学研究的放射自显影方法是敏感和特异的，但在使用放射性同位素时存在固有的缺点。它们还需要耗时和复杂的技术，并且仅为药物在细胞和组织中的定位提供间接检测系统。在一些药物的情况下，自发荧光也被用作标记，但它缺乏敏感性。为了克服这些缺点，并开发一种简单且普遍适用的方法来揭示药物在细胞和组织中的精确定位或摄取，我们建立了目前的免疫细胞化学(ICC)方法。在这项研究中，我们使用了一种蒽环类抗生素柔红霉素(DM)作为原型。在免疫细胞化学…之前，成功的DM ICC需要一系列过程进一步反应：首先用GA固定细胞，然后用NaBH4还原，再用盐酸处理，最后用蛋白酶消化。然后细胞标本与抗DM血清免疫反应，然后用过氧化物酶标记的羊抗兔Ig G/Fab‘，两种免疫试剂中都含有去污剂Triton X-100。本研究成功地在BD细胞的胞核和核周高尔基体染色了DM的所有这些过程，这与DM自发荧光法的结果一致。这种ICC的灵敏度是细胞荧光法的三倍，适用于静脉注射后24小时大鼠肝脏的石蜡切片。注射DM。本研究建立DMD-ICC的原理可应用于其他含有伯氨基(S)的药物。因此，这些用于药物的ICC是直接、准确和简单的新方法，应该有可能用于药物的药理学和毒理学研究，揭示药物在细胞和组织中的定位。较少

A variety of quantitative chemical assays have been used for the pharmacological study of drugs mainly using urine, blood and tissue homogenates from animals. Also, Autoradiography methods, which are commonly employed for pharmacological studies on drugs, are sensitive and specific, but possess drawbacks inherent in the use of a radioisotope. They also require time-consuming and complicated techniques, and only provide an indirect detection system for drug localization in cells and tissues. Autofluorescence is also used as a marker in the case of some drugs, but it is lacking in sensitivity. In order to overcome each of these drawbacks and develop an easy and generally applicable method revealing a precise localization or uptake of the drug in the cells and tissues, we established the present immunocytochemistry (ICC) method. In this study we used an anthracycline antibiotic daunomycin (DM) as a prototype. Successful DM ICC required a series of processes prior to the immunocytochemical … More reaction : the cells were first fixed with GA, then reducted with NaBH4, treated with hydrochloric acid, and finally digested with protease. The cell specimens were then subjected to immunoreaction with anti-DM serum followed by peroxidase-labeled goat anti-rabbit IgG/Fab', and in both the immune reagents the detergent Triton X-100 was contained as well. The present ICC covering all these processes successfully stained for DM in the nucleus and in the perinuclear Golgi region of the cytoplasm of the BD cells, this being consistent with the results obtained by the DM autofluorescence method. This ICC was found to be three times as sensitive as the cytofluorometric method, and applicable to the paraffin sections of the liver of rats 24 hr after an i.v. injection of DM. The principle used in the present study for developing DMD ICC might be applied to other drugs containing the primary amino group(s) in the molecule. Thus, these ICCs for drugs are direct, precise and easy new methods which should have potential for pharmacology and toxicolog studies of drugs, revealing the localization of a drug in cells and tissues. Less