Miloud Tewfik, Henrich Carmen, Hämmerling Günter J
German Cancer Research Center, Department of Molecular Immunology, 69120 Heidelberg, Germany.
J Biomed Opt. 2007 Sep-Oct;12(5):054018. doi: 10.1117/1.2800386.
For bioluminescence imaging (BLI) of small animals, the most commonly used luciferase is Fluc from the firefly, but recently, green (CBGr99) and red (CBRed) click beetle luciferases became available. Because signal attenuation by tissues is lower for red light, red luciferases appear to be advantageous for BLI, but this has not been thoroughly tested. We compare different luciferases for BLI. For this purpose, cell transfectants are generated expressing comparable amounts of CBGr99, CBRed, or Fluc. This is achieved by coexpression of the luciferase with eGFP using the bicistronic 2A system, which results in stoichiometric coexpression of the respective proteins. In vitro, the CBGr99 transfectant exhibits the strongest total photon yield. For in vivo BLI, the transfectants are injected into mice at different locations. At a subcutaneous position, CBGr99 is clearly superior to the other luciferases. When the tumor cells are located in the peritoneum or lung, where more absorption by tissue occurs, CBGr99 and CBRed transfected cells emit a comparable number of red photons and are superior to Fluc, but CBGr99 reaches the maximum of the light emission faster than CBRed. Thus, although CBGr99 emits mainly green light, the high yield of total and red photons makes it an excellent candidate for BLI.
对于小动物的生物发光成像（BLI），最常用的荧光素酶是来自萤火虫的Fluc，但最近，绿色（CBGr99）和红色（CBRed）叩头虫荧光素酶也已可用。由于红光在组织中的信号衰减较低，红色荧光素酶似乎对BLI具有优势，但这尚未得到充分测试。我们比较了用于BLI的不同荧光素酶。为此，生成表达相当数量的CBGr99、CBRed或Fluc的细胞转染子。这是通过使用双顺反子2A系统将荧光素酶与eGFP共表达来实现的，该系统导致相应蛋白质的化学计量共表达。在体外，CBGr99转染子表现出最强的总光子产量。对于体内BLI，将转染子注射到小鼠的不同部位。在皮下位置，CBGr99明显优于其他荧光素酶。当肿瘤细胞位于腹膜或肺部，组织吸收更多的地方时，CBGr99和CBRed转染的细胞发出相当数量的红色光子，并且优于Fluc，但CBGr99比CBRed更快达到发光最大值。因此，尽管CBGr99主要发出绿光，但其高产量的总光子和红色光子使其成为BLI的优秀候选者。
