Experimental Therapeutics and Molecular Imaging Laboratory, Neuroscience Center, Department of Neurology, Massachusetts General Hospital, Charlestown, Massachusetts 02129, United States.
Anal Chem. 2012 Jan 17;84(2):1189-92. doi: 10.1021/ac202833r. Epub 2011 Dec 27.
Secreted Gaussia luciferase (Gluc) has been shown to be a useful tool for ex vivo monitoring of in vivo biological processes. The Gluc level in the blood was used to detect tumor growth, metastasis and response to therapy, gene transfer, and circulating cells viability, as well as transcription factors activation, complementing in vivo bioluminescence imaging. The sensitivity of the Gluc blood assay is limited due to the absorption of blue light by pigmented molecules such as hemoglobin, resulting in quenching of the signal and therefore lower sensitivity. To overcome this problem, we designed an alternative microtiter well-based binding assay in which Gluc is captured first from blood using a specific antibody followed by the addition of coelenterazine and signal acquisition using a luminometer. This assay showed to be over 1 order of magnitude more sensitive in detecting Gluc in the blood as compared to the direct Gluc blood assay enhancing ex vivo monitoring of biological processes.
分泌型海肾荧光素酶（Gluc）已被证明是用于监测体内生物过程的有用工具。血液中的 Gluc 水平可用于检测肿瘤生长、转移和对治疗的反应、基因转移以及循环细胞的活力，还可用于检测转录因子的激活，从而补充体内生物发光成像。由于血红蛋白等色素分子对蓝光的吸收，Gluc 血液检测的灵敏度受到限制，导致信号猝灭，因此灵敏度降低。为了克服这个问题，我们设计了一种替代的微孔板结合测定法，该方法首先使用特异性抗体从血液中捕获 Gluc，然后加入腔肠素并使用发光计进行信号采集。与直接 Gluc 血液测定法相比，该测定法在检测血液中的 Gluc 时灵敏度提高了 1 个数量级以上，从而增强了对生物过程的体外监测。
