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体外培养的细胞不仅可用于研究其形态和机能的变化,而且可进行不同药物学作用时细胞内抗原成分的定位检测及细胞特征的研究。通常盖玻片放入培养皿中作为培养细胞的支持物,细胞在盖玻片上长成单层,可立即固定、染色,进行光学显微镜的观察。在实验中我们发现,加盖玻片的单层细胞培养法也可以直接应用于免疫电镜的包埋前和包埋后染色方法。该技术具有以下优点。(1)使用盖玻片可以避免免疫电镜标本制作过程中,为了搜集细胞而进行的数次刮取、离心。免疫反应及后固定、脱水、浸透、包埋均在盖玻片上进行,使培养细胞的损伤降低至最小程度。(2)如果在直径3.5cm的培养皿中进行免疫过氧化物酶反应大约需要至少200μl抗体,而使用盖玻片法,每张盖玻片则仅需要10μl抗体,大大节约了免疫反应的抗体量,既简便又经济。(3)盖玻片法使包埋后培养细胞呈单层位于包埋块的表面,在每一张超薄切片中都能观察到很多细胞。
In vitro cultured cells can not only be used to study the changes of their morphology and function, but also can be used to study the localization of intracellular antigen components and cell characteristics under different pharmacological effects. The coverslip is usually placed in a petri dish as a support for the cultured cells. The cells grow into a monolayer on the coverslip and can be immediately fixed, stained and observed under an optical microscope. In the experiment, we found that the single-layer cell culture with glass coverslips can also be directly applied to the immunoelectron microscopy before and after embedding staining method. This technique has the following advantages. (1) The use of coverslips can avoid several times of scraping and centrifuging in order to collect cells during the production of immunoelectron microscopy specimens. Immune response and post-fixation, dehydration, soaking, embedding are carried out in the coverslip, so that cultured cells to minimize damage. (2) If the immunoperoxidase reaction in a 3.5 cm diameter Petri dish requires at least 200 μl of antibody, the cover glass method requires only 10 μl of antibody per cover glass, significantly reducing the immune response to antibodies The amount of both simple and economical. (3) The cover glass method enables the cultured cells to be embedded in the monolayer on the surface of the embedded block, and many cells can be observed in each ultrathin section.