3D TIRF microscopy: Combination of the total internal reflection fluorescence microscopy and ultramicrotomy techniques

IF 3.5 2区工程技术 Q2 OPTICS

Optics and Lasers in Engineering Pub Date : 2025-02-28 DOI:10.1016/j.optlaseng.2025.108891

Konstantin E. Mochalov , Olga I. Agapova , Igor I. Agapov , Denis S. Korzhov , Daria O. Solovyeva , Svetlana V. Sizova , Milena S. Shestopalova , Vladimir A. Oleinikov , Anton E. Efimov

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引用次数: 0

Abstract

Improvement of the existing methods of ultrahigh-resolution microscopy and development of new ones, in particular, methods of three-dimensional (3D) analysis and imaging of the ultrastructure of biological objects and nanomaterials, are extremely important tasks in biomedicine, cell biology, and nanotechnology. Here, a method of 3D total internal reflection fluorescence (TIRF) microscopy is proposed that combines ultramicrotomy (successive ultrathin sections of the sample) and TIRF microscopy of the sample surface. The key feature of this combined technique is the use of a specialized diamond ultramicrotome knife both as a tool for making the sections and as a TIRF prism for obtaining layer-by-layer TIRF images within a single measurement procedure. This makes it possible to reconstruct the 3D ultrastructure of objects.

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三维TIRF显微镜：全内反射荧光显微镜和超微切片技术的结合

对现有的超高分辨率显微技术进行改进和开发新的方法，特别是对生物物体和纳米材料的超微结构进行三维分析和成像的方法，是生物医学、细胞生物学和纳米技术的重要任务。本文提出了一种三维全内反射荧光（TIRF）显微镜方法，该方法结合了超显微术（样品的连续超薄切片）和样品表面的TIRF显微镜。这种组合技术的关键特点是使用专门的金刚石超显微刀作为制作切片的工具和在单一测量程序中获得逐层TIRF图像的TIRF棱镜。这使得重建物体的三维超微结构成为可能。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Optics and Lasers in Engineering 工程技术-光学

CiteScore

8.90

自引率

8.70%

发文量

384

审稿时长

42 days

期刊介绍： Optics and Lasers in Engineering aims at providing an international forum for the interchange of information on the development of optical techniques and laser technology in engineering. Emphasis is placed on contributions targeted at the practical use of methods and devices, the development and enhancement of solutions and new theoretical concepts for experimental methods. Optics and Lasers in Engineering reflects the main areas in which optical methods are being used and developed for an engineering environment. Manuscripts should offer clear evidence of novelty and significance. Papers focusing on parameter optimization or computational issues are not suitable. Similarly, papers focussed on an application rather than the optical method fall outside the journal''s scope. The scope of the journal is defined to include the following: -Optical Metrology- Optical Methods for 3D visualization and virtual engineering- Optical Techniques for Microsystems- Imaging, Microscopy and Adaptive Optics- Computational Imaging- Laser methods in manufacturing- Integrated optical and photonic sensors- Optics and Photonics in Life Science- Hyperspectral and spectroscopic methods- Infrared and Terahertz techniques