Division of Ultrastructural Cell Biology, Department of Anatomy, University of Miyazaki, Miyazaki, Japan.
Hitachi High-Tech Corporation, Tokyo, Japan.
Commun Biol. 2021 Jun 10;4(1):710. doi: 10.1038/s42003-021-02246-3.
Immunocytochemistry visualizes the exact spatial location of target molecules. The most common strategy for ultrastructural immunocytochemistry is the conjugation of nanogold particles to antibodies as probes. However, conventional nanogold labelling requires time-consuming nanogold probe preparation and ultrathin sectioning of cell/tissue samples. Here, we introduce an in situ strategy involving nanogold nucleation in immunoenzymatic products on universal paraffin/cryostat sections and provide unique insight into nanogold development under hot-humid air conditions. Nanogold particles were specifically localized on kidney podocytes to target synaptopodin. Transmission electron microscopy revealed secondary growth and self-assembly that could be experimentally controlled by bovine serum albumin stabilization and phosphate-buffered saline acceleration. Valuable retrospective nanogold labelling for gastric H/K-ATPase was achieved on vintage immunoenzymatic deposits after a long lapse of 15 years (i.e., 15-year-old deposits). The present in situ nanogold labelling is anticipated to fill the gap between light and electron microscopy to correlate cell/tissue structure and function.
免疫细胞化学可直观呈现靶分子的确切空间位置。超微结构免疫细胞化学最常用的策略是将纳米金颗粒与抗体偶联作为探针。然而，传统的纳米金标记需要耗时的纳米金探针制备和细胞/组织样品的超薄切片。在这里，我们引入了一种原位策略，即在石蜡/冰冻切片的免疫酶产物中进行纳米金成核，并提供了在湿热空气条件下纳米金发展的独特见解。纳米金颗粒特异性地定位于肾脏足细胞以靶向突触蛋白。透射电子显微镜显示，通过牛血清白蛋白稳定和磷酸盐缓冲盐水加速，可以对二次生长和自组装进行实验控制。通过在长时间（即 15 年）后对陈旧的免疫酶沉积物进行回溯性纳米金标记，成功实现了对胃 H/K-ATPase 的有价值的纳米金标记。目前的原位纳米金标记有望填补光镜和电镜之间的空白，以关联细胞/组织结构和功能。
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