2025-06-18 順天堂大学
図1:ITGAM POD-nAbの立体構造予測
本論文で作成したPOD-nAbの一つであるITGAM POD-nAbの立体構造予測。ITGAM POD-nAbは脳の免疫細胞であるミクログリア表面に発現する抗原を認識する。POD-nAbは抗原を認識するナノボディ(図中左側)と高感度検出を可能にするペルオキシダーゼ(図中右側)の融合タンパクをして調製される。立体構造はAlphaFold3*10により予測した。
<関連情報>
ペルオキシダーゼ標識ナノボディとFT-GOシグナルチラミド増幅法を用いた三次元免疫組織化学法の開発 A three dimensional immunolabeling method with peroxidase-fused nanobodies and fluorochromized tyramide-glucose oxidase signal amplification
Kenta Yamauchi,Masato Koike & Hiroyuki Hioki
Communications Biology Published:18 June 2025
DOI:https://doi.org/10.1038/s42003-025-08317-z
Abstract
Three dimensional immunohistochemistry (3D-IHC), immunolabeling of 3D tissues, reveals the spatial organization of molecular and cellular assemblies in the context of the tissue architecture. Deep and rapid penetration of antibodies into 3D tissues and highly sensitive detection are critical for high-throughput imaging analysis of immunolabeled 3D tissues. Here, we report a nanobody (nAb)-based 3D-IHC, POD-nAb/FT-GO 3D-IHC, for high-speed and high-sensitive detection of targets within 3D tissues. Peroxidase-fused nAbs (POD-nAbs) enhanced immunolabeling depth and allowed for highly sensitive detection by combined with a fluorescent tyramide signal amplification system, Fluorochromized Tyramide-Glucose Oxidase (FT-GO). Multiplex labeling was implemented to the 3D-IHC by quenching POD with sodium azide. Using the 3D-IHC technique, we successfully visualized somata and processes of neuronal and glial cells in millimeter-thick mouse brain tissues within three days. Given its high-speed and high-sensitive detection, our 3D-IHC protocol, POD-nAb/FT-GO 3D-IHC, would provide a useful platform for histochemical analysis in 3D tissues.
