Near‐Infrared Bioluminescence Imaging with a through‐Bond Energy Transfer Cassette

Abstract

A coelenterazine (CTZ) analogue emitting near‐infrared (NIR) bioluminescence was synthesized for through‐bond energy transfer (TBET)‐based imaging modalities. The analogue, named Cy5‐CTZ, was prepared by conjugating cyanine‐5 (Cy5) dye to CTZ through an acetylene linker. This novel derivative is intrinsically fluorescent and emits NIR‐shifted luminescence upon reacting with an appropriate luciferase, the Renilla luciferase. This Cy5‐CTZ substrate is optically stable in physiological samples and rapidly permeabilize through the plasma membrane into the cytosolic compartment of live cells.

Near-infrared fluorescent protein and bioluminescence-based probes for high-resolution in vivo optical imaging

“…BRET is a special case of resonance energy transfer, where the donor is a luciferase/luciferin system. As BRET acceptors, fluorescent materials such as FPs, 14,84,85 organic dyes, 35,[86][87][88][89][90][91] and QDs have been employed (Fig. 5a-c).…”

“…To date, several types of far-red and NIR fluorescent dyes have been employed as BRET acceptors. 27,36,[86][87][88][89][90][91] To induce efficient energy transfer in the BRET system of a luciferase-dye conjugate, the bioluminescence spectrum (donor) of luciferin should be overlapped with the absorption spectrum of the fluorescence dye (acceptor). High fluorescence quantum yields as well as high extinction coefficients for the acceptor dyes are also important to achieve efficient BRET, leading to strong NIR emission from the NIR acceptor dye.…”

“…They developed BRET based NIR imaging using the CTZ-Cy5 substrate as a through-bond energy transfer cassette. 87 3.2.3. BRET probes using NIR QDs.…”

Palette of Luciferases: Natural Biotools for New Applications in Biomedicine

“…To overcome these limitations, a number of RLuc-based reporters with improved properties have been obtained using random or site-directed mutagenesis, such as mutants with increased resistance to inactivation by blood serum, enhanced brightness, or proteins with red-shifted spectra [ 44 , 47 , 49 , 49 ]. To expand the scope of applications of RLuc, several coelenterazine analogs with increased brightness of luminescence and red-shifted emission have also been generated [ 49 , 50 ].…”

Palette of Luciferases: Natural Biotools for New Applications in Biomedicine

“…Для преодоления раз-личных ограничений методами случайного или сайтнаправленного мутагенеза получен ряд репортеров на основе RLuc с улучшенными свойствами: повышенной устойчивостью к инактивации сывороткой крови, повышенной яркостью, а также белки со спектрами, смещенными в длинноволновую область [44,[47][48][49]. Чтобы расширить сферу применения RLuc, разработаны более яркие аналоги целентеразина с красным спектром люминесценции [49,50].…”