Emission spectra of bioluminescent reporters and interaction with mammalian tissue determine the sensitivity of detection in vivo -: art. no. 041210
Emission spectra of bioluminescent reporters and interaction with mammalian tissue determine the sensitivity of detection in vivo -: art. no. 041210
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DOI：
10.1117/1.2032388
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发表时间：
2005-07-01
影响因子：
3.5
通讯作者：
Contag, CH
中科院分区：
文献类型：
--
作者：
Zhao, H;Doyle, TC;Contag, CH
体内生物发光成像依赖于由体内的生物发光酶发射的光克服组织衰减的影响。理解这种关系对于信号的检测和定量是必不可少的。我们研究了四种具有不同发射光谱的密码子优化的酶，包括来自萤火虫（FLuc）、点击甲虫（CBGr 68、CBRed）和海肾（Renilla reniformins）（hRLuc）的酶。在25摄氏度，在体外λ（最大值）的这些报告分别为578，543，615和480 nm，在体温，37摄氏度，亮度增加，萤火虫酶表现出34 nm的光谱红移。使用一系列20 nm带通滤波器和冷却的电荷耦合器件（CCD）摄像机，由于组织效应的光谱位移和衰减进行了评价。衰减增加，并且发射光的光谱对于相对于浅表起源源自体内更深处的信号发生红移。来自CBGr 68和hRLuc的信号的组织衰减大于来自Fluc和CBRed的信号的组织衰减。为了进一步探测组织效应，通过CBGr 68和CBRed之间的基因融合产生了宽光谱发射器。这些导致酶具有更宽的发射光谱，具有两个峰，其强度受温度和组织深度的不同影响。这些光谱测量数据允许更好地理解这些报告者如何在体内使用，以及它们可以揭示活体中的生物过程。(C)2005年，由光学仪器工程师学会（Society of Photo-Optical Instrumentation Engineers）主办。
In vivo bioluminescence imaging depends on light emitted by luciferases in the body overcoming the effect of tissue attenuation. Understanding this relationship is essential for detection and quantification of signal. We have studied four codon optimized luciferases with different emission spectra, including enzymes from firefly (FLuc), click beetle (CBGr68, CBRed) and Renilla reniformins (hRLuc). At 25 degrees C, the in vitro lambda(max) of these reporters are 578, 543, 615, and 480 nm, respectively; at body temperature, 37 degrees C, the brightness increases and the firefly enzyme demonstrates a 34-nm spectral red shift. Spectral shifts and attenuation due to tissue effects were evaluated using a series of 20-nm bandpass filters and a cooled charge-coupled device (CCD) camera. Attenuation increased and the spectra of emitted light was red shifted for signals originating from deeper within the body relative to superficial origins. The tissue attenuation of signals from CBGr68 and hRLuc was greater than from those of Fluc and CBRed. To further probe tissue effects, broad spectral emitters were created through gene fusions between CBGr68 and CBRed. These resulted in enzymes with broader emission spectra, featuring two peaks whose intensities are differentially affected by temperature and tissue depth. These spectral measurement data allow for improved understanding of how these reporters can be used in vivo and what they can reveal about biological processes in living subjects. (C) 2005 Society of Photo-Optical Instrumentation Engineers.