Biomedical Applications of High-troughput Raman Flow Cytometry

2019 Fiscal Year Annual Research Report

• Project/Area Number: 19F19805
• Research Institution: The University of Tokyo
• Principal Investigator: 合田 圭介 東京大学, 大学院理学系研究科(理学部), 教授 (70518696)
• Co-Investigator(Kenkyū-buntansha): GALA DE PABLO JULIA 東京大学, 理学(系)研究科(研究院), 外国人特別研究員
• Project Period (FY): 2019-11-08 – 2022-03-31
• Keywords: FT-CARS / Raman Microalgae / Euglena Starch / Carotenoid / Carbohydrate Paramylon

Outline of Annual Research Achievements

The pulse-shaping capabilities were improved and a transform-limited pulse was achieved, improving the sensitivity and signal quality for carbohydrates and carotenoids. Heavy ions mutagenesis was performed in Euglena gracilis, identifying via FT-CARS paramylon hyperaccumulating mutants, whose genetic analysis is being done. Analysis of starch accumulation in Chromochloris zofingiensis via FT-CARS was done and enzymatic analysis is now being done as a golden standard. Sorting of multiple microalgae species was achieved. Human blood signal was obtained via FT-CARS. However, leukocyte detection is still challenging with the current system sensitivity. An additional application of the FT-CARS system was identified as microplastics detection and is being researched.

Current Status of Research Progress

3: Progress in research has been slightly delayed.

Reason

Blood was detected satisfactorily, but as white blood cells could not be detected, the initial project is on hold, and thus other bio-applications are being explored until the sensitivity of the system can be improved. However, satisfactory progress is being done on other applications and the system capabilities have been improved, with the potential to detect mammalian white blood cells by the end of the project.

Strategy for Future Research Activity

During the remaining of my research fellowship I will aim my efforts on 4 projects: i) Directed Evolution of Euglena gracilis: I will focus on characterizing the isolated mutants by looking at their expression of paramylon in high and low carbon media as well as genetic analysis. ii) Otimization of starch and carotenoids expression of Chromochloris zofingiensis and mutagenesis to identify astaxanthin expressing mutants. iii) in-flow FT-CARS detection for microplastics analysis. iv) Increasing sensitivity for leukocyte detection in blood. Additionally, imaging capabilities will be integrated in the system, as well as increasing bandwidth.