基因工程人抗角蛋白抗体在角质形成细胞内的反应定位

Immunohistochemical localization of a genetically engineered human anti - keratin antibody in keratinocytes

目的:探索一株基因工程人源性抗角蛋白Fab抗体在培养的角质形成细胞内的反应定位。方法:利用从噬菌体抗体库中筛选到的特异表达抗角蛋白Fab片段的质粒转化大肠杆菌,IPTG诱导表达出Fab抗体,纯化鉴定后将该抗体作用于培养的人角质形成细胞,在不同浓度和不同的作用时间应用免疫荧光细胞染色法及共聚焦显微镜观察该抗体在角质形成细胞内的反应定位,同时以基因工程抗HBsAg人Fab和1%BSA为对照。结果:基因工程人源性抗角蛋白Fab抗体作用于角质形成细胞的胞浆,呈明显的绿色荧光,着色均匀,细胞核未见着色,两种对照均未见着色。结论:基因工程人源性抗角蛋白Fab抗体,可与培养状态下的角质形成细胞胞浆中其特异性的抗原相结合,这为其临床应用奠定了基础。

Objective: To explore the immunoreactive localization of a genetically engineered human antibody (GEHA) against keratin in keratinocytes. Methods: The specific anti - keratin Fab was previously cloned from a phage antibody library. After being expressed and characterized, the antibodies of different concentration were added to cultured keratinocytes, and the immunoreactive localization in keratinocytes was identified by immunofluorescence and confocal microscopy at different times, taking some other keratinocytes were cultured with a GEHA against HBsAg and 1 % BSA as control. Results: Bright green fluorescence was seen in cytoplasm of keratinocytes cultured with the anti- keratin GEHA. Stains of the keratinocytes were relatively homogeneously. No bright green fluorescence was seen in nuclear of keratinocytes. And no bright green fluorescence was seen in keratinocytes cultured with a GEHA against HBsAg and 1% BSA. Conclusion: The anti - keratin GEHA can penetrate into the cytoplasm of cultured keratinocytes and its immunoreactivity was localized only in cytoplasm. The exploration may lay a role for its clinical application.