新型NanoLuc螢光素酶底物助力動物體內雙色生物發光成像
作者:
小柯機器人發布時間:2020/7/16 16:00:48
2020年7月13日,史丹福大學Michael Z. Lin課題組與Promega Bioscience公司的Thomas A. Kirkland研究團隊在《自然—方法學》雜誌發表論文,報導新型NanoLuc螢光素酶底物,實現小鼠體內明亮two-population生物發光成像。
該研究團隊報導了一種新的底物,hydrofurimazine,其提高的水溶性有利於向小鼠遞送更高劑量。
結合Antares螢光素酶,底物hydrofurimazine在肝臟中表現出和使用AkaLuc螢光素酶及其底物AkaLuc相似的亮度。
進一步的化學探索產生了第二種底物,fluorofurimazine,在小鼠體內具有更高的亮度。
研究人員使用Antares螢光素酶和底物fluorofurimazine追蹤腫瘤大小,並用AkaLuc螢光素酶和底物AkaLumine追蹤CAR-T細胞,實現了在同一隻老鼠上使用兩種螢光素酶系統追蹤監測兩種細胞群體的應用。
Hydrofurimazine和fluorofurimazine作為具有更高溶解度和生物利用度的NanoLuc底物,能大大增強體內生物發光信號,並使高亮度雙色生物發光成像成為可能。
據悉,對於體內兩種生物活動的靈敏檢測一直是生物發光成像的目標。Antares是螢光素酶NanoLuc與橙色螢光蛋白CyOFP的結合體,已成為一種明亮的生物發光報告分子,對螢火蟲螢光素酶(FLuc)及其衍生物(如AkaLuc)具有正交的底物特異性。然而,Antares在老鼠體內的亮度受限於NanoLuc螢光素酶底物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