Department of Bacteriology, The Jikei University School of Medicine, 3-25-8 Nishi-Shimbashi, Minato-ku, Tokyo, 105-8461, Japan.
Jikei Center for Biofilm Science and Technology, The Jikei University School of Medicine, 3-25-8 Nishi-Shimbashi, Minato-ku, Tokyo, 105-8461, Japan.
Commun Biol. 2023 Jan 23;6(1):38. doi: 10.1038/s42003-022-04396-4.
Whole-biofilm imaging at single-cell resolution is necessary for system-level analysis of cellular heterogeneity, identification of key matrix component functions and response to immune cells and antimicrobials. To this end, we developed a whole-biofilm clearing and imaging method, termed instantaneous clearing of biofilm (iCBiofilm). iCBiofilm is a simple, rapid, and efficient method involving the immersion of biofilm samples in a refractive index-matching medium, enabling instant whole-biofilm imaging with confocal laser scanning microscopy. We also developed non-fixing iCBiofilm, enabling live and dynamic imaging of biofilm development and actions of antimicrobials. iCBiofilm is applicable for multicolor imaging of fluorescent proteins, immunostained matrix components, and fluorescence labeled cells in biofilms with a thickness of several hundred micrometers. iCBiofilm is scalable from bacterial to fungal biofilms and can be used to observe biofilm-neutrophil interactions. iCBiofilm therefore represents an important advance for examining the dynamics and functions of biofilms and revisiting bacterial and fungal biofilm formation.
单细胞分辨率全生物膜成像对于细胞异质性的系统分析、关键基质成分功能的鉴定以及对免疫细胞和抗菌药物的反应是必要的。为此，我们开发了一种全生物膜清除和成像方法，称为生物膜瞬时清除（iCBiofilm）。iCBiofilm 是一种简单、快速和有效的方法，包括将生物膜样品浸入折射率匹配介质中，使共焦激光扫描显微镜能够即时对全生物膜进行成像。我们还开发了非固定 iCBiofilm，能够对生物膜发育和抗菌药物作用进行实时和动态成像。iCBiofilm 适用于厚度几百微米的生物膜中荧光蛋白、免疫染色基质成分和荧光标记细胞的多色成像。iCBiofilm 可从细菌生物膜扩展到真菌生物膜，可用于观察生物膜-中性粒细胞相互作用。因此，iCBiofilm 代表了检查生物膜动力学和功能以及重新审视细菌和真菌生物膜形成的重要进展。
