小柯机器人

2020年7月13日，斯坦福大学Michael Z. Lin课题组与Promega Bioscience公司的Thomas A. Kirkland研究团队在《自然—方法学》杂志发表论文，报道新型NanoLuc荧光素酶底物，实现小鼠体内明亮two-population生物发光成像。

该研究团队报道了一种新的底物，hydrofurimazine，其提高的水溶性有利于向小鼠递送更高剂量。

结合Antares荧光素酶，底物hydrofurimazine在肝脏中表现出和使用AkaLuc荧光素酶及其底物AkaLuc相似的亮度。

进一步的化学探索产生了第二种底物，fluorofurimazine，在小鼠体内具有更高的亮度。

研究人员使用Antares荧光素酶和底物fluorofurimazine追踪肿瘤大小，并用AkaLuc荧光素酶和底物AkaLumine追踪CAR-T细胞，实现了在同一只老鼠上使用两种荧光素酶系统追踪监测两种细胞群体的应用。

Hydrofurimazine和fluorofurimazine作为具有更高溶解度和生物利用度的NanoLuc底物，能大大增强体内生物发光信号，并使高亮度双色生物发光成像成为可能。

据悉，对于体内两种生物活动的灵敏检测一直是生物发光成像的目标。Antares是萤光素酶NanoLuc与橙色荧光蛋白CyOFP的结合体，已成为一种明亮的生物发光报告分子，对萤火虫萤光素酶（FLuc）及其衍生物（如AkaLuc）具有正交的底物特异性。然而，Antares在老鼠体内的亮度受限于NanoLuc荧光素酶底物furimazine的低溶解性和低生物利用度。

附：英文原文

Title: Novel NanoLuc substrates enable bright two-population bioluminescence imaging in animals

Author: Yichi Su, Joel R. Walker, Yunhee Park, Thomas P. Smith, Lan Xiang Liu, Mary P. Hall, Louai Labanieh, Robin Hurst, David C. Wang, Lance P. Encell, Namdoo Kim, Feijie Zhang, Mark A. Kay, Kerriann M. Casey, Robbie G. Majzner, Jennifer R. Cochran, Crystal L. Mackall, Thomas A. Kirkland, Michael Z. Lin

Issue&Volume: 2020-07-13

Abstract: Sensitive detection of two biological events in vivo has long been a goal in bioluminescence imaging. Antares, a fusion of the luciferase NanoLuc to the orange fluorescent protein CyOFP, has emerged as a bright bioluminescent reporter with orthogonal substrate specificity to firefly luciferase (FLuc) and its derivatives such as AkaLuc. However, the brightness of Antares in mice is limited by the poor solubility and bioavailability of the NanoLuc substrate furimazine. Here, we report a new substrate, hydrofurimazine, whose enhanced aqueous solubility allows delivery of higher doses to mice. In the liver, Antares with hydrofurimazine exhibited similar brightness to AkaLuc with its substrate AkaLumine. Further chemical exploration generated a second substrate, fluorofurimazine, with even higher brightness in vivo. We used Antares with fluorofurimazine to track tumor size and AkaLuc with AkaLumine to visualize CAR-T cells within the same mice, demonstrating the ability to perform two-population imaging with these two luciferase systems. NanoLuc substrates with improved solubility and bioavailability, hydrofurimazine and fluorofurimazine, strongly enhance bioluminescence signals in vivo and enable bright dual-color bioluminescent imaging with AkaLuc and AkaLumine.

DOI: 10.1038/s41592-020-0889-6

Source: https://www.nature.com/articles/s41592-020-0889-6

Nature Methods：《自然—方法学》，创刊于2004年。隶属于施普林格·自然出版集团，最新IF：47.99
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