粉尘螨2型变应原抗原定位的研究

Study on the Localization of a Major Allergen Der f 2 by Fluorescence Immunohistochemical Method

目的研究粉尘螨主要变应原Der f 2在螨体内的定位。方法采用免疫荧光抗原定位观察。先制备Der f 2单克隆抗体,再制作粉尘螨石蜡切片。切片经二甲苯脱蜡及梯度乙醇脱水,入3%H2O2孵育10 min,PBS漂洗后依次加入单克隆抗体血清37℃孵育1 h、生物素标记的羊抗鼠IgG抗体(1∶100)室温孵育60 min、抗生物素蛋白-生物素过氧化物酶-异硫氰酸荧光素复合物(SABC-FITC)(1∶100)室温孵育60 min。PBS漂洗,伊文氏蓝处理切片5min,最后丙三醇封片。用抗重组Der f2单克隆抗体作一抗孵育,再选用抗鼠IgG荧光抗体为二抗,经荧光染色,观察Der f2在螨体内的位置。用正常小鼠血清(1∶100)代替小鼠单克隆抗体血清作为阴性对照。结果粉尘螨Der f2在中肠组织(包括中肠和后肠)和肠内容物均显示较强阳性,特别是肠内容物更明显。结论Der f 2分布于粉尘螨的中肠组织及其肠内容物。

Objective To localize Der f 2 in the body of Dermatophogoides farinae. Methods Live mites were embedded. Serial mite paraffin sections were made and checked under light microscope. The sections were incubated with anti-recombinant Der f 2 monoclonal antibody as the first antibody. They were then reacted with the fluorescent isothiocyanate conjugated goat anti-mouse IgG as the secondary antibody. The sections were examined with fluorescent light microscopy. Results Der f 2 reacting with immunofluorescent antibodies was found localized in the tissue and contents of the mid-gut of the mite. Conclusion The major allergen Der f 2 distributes in the gut and fecal pellets of Dermatophogoides farinae.