Efficient targeting method for correlative light and electron microscopy

• Project/Area Number: 22K20741
• Research Category: Grant-in-Aid for Research Activity Start-up
• Allocation Type: Multi-year Fund
• Review Section: 0802:Biomedical structure and function and related fields
• Research Institution: Yamaguchi University
• Principal Investigator: Yoshimura Azumi 山口大学, 大学院医学系研究科, 講師 (90467340)
• Project Period (FY): 2022-08-31 – 2024-03-31
• Project Status: Completed (Fiscal Year 2023)
• Budget Amount: ¥2,860,000 (Direct Cost: ¥2,200,000、Indirect Cost: ¥660,000); Fiscal Year 2023: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2022: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
• Keywords: 電子顕微鏡 / 光-電子相関顕微鏡法

Outline of Research at the Start

光-電子相関顕微鏡法は分子の位置情報と微細形態の相関を可能にする重要な手法であるが、例えば組織などの大きな試料においては蛍光標識した微小な領域を電子顕微鏡試料中で探し出すことは容易ではない。本研究は、光学顕微鏡観察によって特定した試料中の目的領域の微細形態を効率的に取得するための3次元ターゲティング法の開発を目指す。

Outline of Final Research Achievements

Correlative light and electron microscopy (CLEM) enables correlation of molecular and structural information and allows qualitative and quantitative analysis of regions of interest (ROI) specified by light microscopy. The correlation however becomes challenging as the specimen size increases, due to the difficulty in ROI targeting after EM sample preparation. This study aims to develop an efficient ROI targeting method for CLEM that can be performed with widely used fluorescence microscopy. Using a fluorescent label detectable both before and after EM samples preparation, landmark structures were introduced. Based on their positions detected by fluorescence microscopy at different steps of the sample preparation, step-wise ROI targeting was achieved.

Academic Significance and Societal Importance of the Research Achievements

立体的に大きな試料における光-電子相関顕微鏡法（CLEM）には、目的領域のターゲティングに先端的設備を要する、電子観察のデータサイズが増大しスループットが低くなるなどの問題点がある。本課題で実現されたターゲティング法は、一般的な蛍光観察のみを用いて実行可能なもので、手法の簡略化、自動化を図り汎用性を高めることで、3次元でのCLEMを効率化することが期待される。