Pâques M, Teppema J S, Beuvery E C, Abdillahi H, Poolman J T, Verkleij A J
National Institute of Public Health and Environmental Protection, Bilthoven, The Netherlands.
Infect Immun. 1989 Feb;57(2):582-9. doi: 10.1128/iai.57.2.582-589.1989.
The parallel application of two electron microscopic immunogold labeling procedures was used to assess the surface exposure and accessibility of gonococcal and meningococcal surface antigens. Monoclonal antibodies were used as markers for the surface antigens, i.e., outer membrane proteins and lipooligosaccharides. To evaluate the labeling densities obtained after incubation of whole bacteria in suspension or ultrathin cryosections of bacteria, a method of electron microscopic quantitation was developed. Incubation of whole bacterial suspensions with monoclonal antibodies and protein A-gold resulted in specific labeling of the bacterial surfaces. However, the labeling densities varied largely in each cell. By contrast, cryosections showed uniform heavy labeling densities at the surface of the outer membranes of all cells. Apparently, by sectioning the cells the antigen-masking barrier could be evaded, and steric hindrance was no longer restrictive. Thus, a better estimate of both the presence and the surface exposure, i.e., the accessibility of antigens, could be made. Such information is essential for us to better understand host-bacterial interactions and to develop new vaccines.
运用两种电子显微镜免疫金标记程序并行的方法，来评估淋球菌和脑膜炎球菌表面抗原的表面暴露情况及可及性。单克隆抗体被用作表面抗原的标志物，即外膜蛋白和脂寡糖。为了评估在悬浮状态下对完整细菌进行孵育或对细菌超薄冰冻切片进行孵育后获得的标记密度，开发了一种电子显微镜定量方法。用单克隆抗体和蛋白A-金对完整细菌悬液进行孵育，可使细菌表面产生特异性标记。然而，每个细胞中的标记密度差异很大。相比之下，冰冻切片显示所有细胞外膜表面的标记密度均一且较高。显然，通过对细胞进行切片，可以避开抗原屏蔽屏障，空间位阻也不再具有限制作用。因此，可以更好地估计抗原的存在情况以及表面暴露情况，即抗原的可及性。这些信息对于我们更好地理解宿主与细菌的相互作用以及开发新型疫苗至关重要。
