Development of a generalizable toolkit for rapidly improving the sensitivity of fluorescent immunosensor proteins
| Project/Area Number |
21K14468
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| Research Category |
Grant-in-Aid for Early-Career Scientists
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| Allocation Type | Multi-year Fund |
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| Review Section |
Basic Section 27040:Biofunction and bioprocess engineering-related
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| Research Institution | Tokyo Institute of Technology |
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Principal Investigator |
朱 博 東京工業大学, 科学技術創成研究院, 助教 (70886605)
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| Project Period (FY) |
2021-04-01 – 2024-03-31
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| Project Status |
Granted (Fiscal Year 2022)
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| Budget Amount *help |
¥3,510,000 (Direct Cost: ¥2,700,000、Indirect Cost: ¥810,000)
Fiscal Year 2022: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2021: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
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| Keywords | Immunosensor / mRNA display / Quenchbody / SARS-CoV-2 / Phage display / NGS / バイオテクノロジー / タンパク質 / バイオセンサー / 蛍光 / mRNA-display |
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| Outline of Research at the Start |
Quenchbody（Q-body）は, 抗体あるいはその断片を部位特異的に蛍光修飾して得られる, 蛍光免疫センサーである. Q-bodyは, クエンチ解消原理に基づき様々な抗原を迅速・簡便に検出可能と考えられている. しかし, 現状では, 一つの抗原が一つのQ-bodyの蛍光色素を光らせる事しかできず, Q-bodyの感度は, 親抗体の動力学的特性によって制限されると考えられる. 本研究では, Q-bodyの感度を短期間で向上させるためのツールキットを開発することを目的とする. このツールキットにより, Q-bodyに基づく免疫測定法の開発期間を短縮し, 緊急事態への迅速な対応を可能にする.
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| Outline of Annual Research Achievements |
The research project is about making a toolkit for improving the performance of Quenchbody-based immunosensor (Quenchbody) in a short period. It will increase the feasibility of the quenchbody while responding to the needs of society in a fast manner, such as combating pandemics.
In the second year, the mRNA-display procedures were extensively optimized to get a functional and large Quenchbody for the linker screening. The cell-free synthesized Bone Gla protein (BGP) antibody fragment was successfully labeled with a fluorophore in an mRNA-display-required working procedure. The affinity and immunosensor function of the labeled antibody were confirmed. In addition, a phage-display-based Quenchbody library was created with unexpectedly good feasibility for functional linker sequence enrichment. Two rounds of selection of a BGP Quenchbody library with the randomized fluorophore linker sequences were performed and a clear motif was enriched, which was confirmed via next-generation sequencing.
In the meanwhile, the crowding agent function for rapidly improving the Quenchbody function was fully tested on a SARS-CoV-2 Quenchbody. The related results have been published in an International journal and the paper was selected as HOT Article and a Back Cover.
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| Current Status of Research Progress |
Current Status of Research Progress
3: Progress in research has been slightly delayed.
Reason
The research progress was slightly affected by the COVID-19 pandemic. An alternative phage display-based Quenchbody library was successfully created with unexpectedly better feasibility to screen linker sequences. Therefore, additional experiment is needed to finalize the rest of the screening and analysis for a higher grade outcome.
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| Strategy for Future Research Activity |
The mRNA-display procedures for Quenchbody will be further optimized for producing a useful Quenchbody library. The alternative and robust Quenchbody-displayed phage library will be screened to find better linker sequences for improved response and signal amplification. One of the targeting antibodies will be shifted from the cortisol antibody to another biomarker thyroxine with better affinity. Instead of the linear model, a machine learning-based model will be generated to predict the function of the linkers by using the next-generation sequence datasets. The related output will be organized as another research article for publication and conference presentation.
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Report
(2 results)
Research Products
(22 results)
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[Journal Article] The Patrol Yeast: A new biosensor armed with antibody-receptor chimera detecting a range of toxic substances associated with food poisoning2023
Author(s)
Jiulong Su, Bo Zhu, Akihito Inoue, Hiroyuki Oyama, Izumi Morita, Jinhua Dong, Takanobu Yasuda, Yoshiko Sugita-Konishi, Tetsuya Kitaguchi, Norihiro Kobayashi, Shiro Miyake, Hiroshi Ueda
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Journal Title
Biosensors and Bioelectronics
Volume: 219
Pages: 114793-114793
DOI
Related Report
Peer Reviewed / Open Access / Int'l Joint Research
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