Significant advances have occurred in the area of high-resolution scanning electron microscopy (SEM), especially related to methodologies that allow the observation of intracellular structures that are exposed either by successive abrasion with a gallium ion beam or by sectioning in epoxy-embedded cells. Images of series of successively exposed surfaces can then be rendered into 3D models. Here, we report our observations by combining this approach with classical cytochemical methods to facilitate the 3D reconstruction of labeled structures and organelles. We used epimastigotes of Trypanosoma cruzi whose endocytic pathway was labeled with horseradish peroxidase, followed by fixation and detection of the peroxidase activity using the classical diaminobenzidine-osmium method followed by incubation with thiocarbohydrazide, which increases the concentration of osmium at the sites where the enzyme is located as well as the contrast of lipid-containing structures. This procedure allows not only a better visualization of membranous structures and lipid inclusions but can also easily identify the endocytic tracer (HRP) inside the cell. All structures involved in the endocytic activity could be traced and reconstructed.

高分辨率扫描电子显微镜（SEM）领域取得了重大进展，特别是与允许观察细胞内结构的方法学有关，这些细胞结构是通过连续磨蚀镓离子束或通过切入环氧包埋的细胞而暴露的。然后可以将一系列连续暴露的表面的图像渲染为3D模型。在这里，我们通过将这种方法与经典细胞化学方法相结合来报告我们的观察结果，以促进标记结构和细胞器的3D重建。我们使用了锥虫锥虫其内吞途径用辣根过氧化物酶标记，然后使用经典的二氨基联苯胺-法固定和检测过氧化物酶活性，然后与硫代碳酰肼一起孵育，这会增加酶所在位点的os浓度以及对比含脂质的结构。此过程不仅可以更好地可视化膜结构和脂质包裹体，还可以轻松识别细胞内的内吞示踪剂（HRP）。可以追踪和重建参与胞吞活性的所有结构。