牙本质内Ⅰ型胶原纤维形态的免疫标记初探

Preliminary research on immune labelling of typeⅠ collagen fibril in dentin

目的初步探讨进行免疫标记的牙本质脱矿后暴露的Ⅰ型胶原纤维。方法利用免疫组化技术和场发射扫描电镜(FEISEM)相结合,对暴露胶原纤维的形态以及结构完整性进行评价。结果通过FEISEM观察,牙本质脱矿后暴露的Ⅰ型胶原纤维,可以结合抗Ⅰ型胶原单克隆抗体以及胶体金标记的相应二抗,胶体金标记颗粒分布均匀,可与暴露的胶原纤维上的抗原表位相结合,实现免疫定位。结论免疫组化技术和FEISEM相结合,可以直观观察胶原纤维立体网络的超微结构,是一种准确的、可复制的、可行的方法 。

Objective This study was designed to evaluate the morphology of type Ⅰ collagen in demineralized dentin using an immunocytochemical technique. Methods After dentin was etched with 37% phosphoric acid gel for 15 s, a double immune labeling procedure was applied, and then the morphological and structural integrity evaluation of exposed collagen fibril was carried out under a field emission in-lens scanning electron microscope （FEISEM）. Results Type Ⅰ collagen was labeled with secondary antibodies conjugated with 20 nm colloidal gold particles. The intense and uniform labeling signal was detected on the major fibrils indicating that the anti-type Ⅰ collagen antibody could bind with favorable interaction sites on the exposed type Ⅰ collagen molecular surface. Conchtsions The ultrastructure of three-dimensional collagen fibril network could be identified by immunocytochemistry technology under FEISEM observation, which represented an accurate, reproducible and feasible method.