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Bioluminescence Detection of Superoxide Anion Using Aequorin.
Analytical Chemistry ( IF 6.7 ) Pub Date : 2019-10-02 , DOI: 10.1021/acs.analchem.9b02293 Hossein Rahmani 1 , Fahimeh Ghavamipour 1 , Reza H Sajedi 1
Analytical Chemistry ( IF 6.7 ) Pub Date : 2019-10-02 , DOI: 10.1021/acs.analchem.9b02293 Hossein Rahmani 1 , Fahimeh Ghavamipour 1 , Reza H Sajedi 1
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Although the superoxide anion (O2–·) is generated during normal cellular respiration and has fundamental roles in a wide range of cellular processes, such as cell proliferation, migration, apoptosis, and homeostasis, its dysregulation is associated with a variety of diseases. Regarding these prominent roles in biological systems, the development of accurate methods for quantification of superoxide anion has attracted tremendous research attention. Here, we evaluated aequorin, a calcium-dependent photoprotein, as a potential bioluminescent reporter protein of superoxide anion. The mechanism is based on the measurement of aequorin bioluminescence, where the lower the concentration of coelenterazine under the oxidation of superoxide anion, the lower the amount aequorin regeneration, leading to a decrease in bioluminescence. The bioluminescence intensity of aequorin was proportional to the concentration of superoxide anion in the range from 4 to 40 000 pM with a detection limit (S/N = 3) of 1.2 pM, which was 5000-fold lower than those of the chemiluminescence methods. The proposed method exhibited high sensitivity and has been successfully applied to the determination of superoxide anion in the plant cell samples. The results could suggest a photoprotein-based bioluminescence system as a highly sensitive, specific, and simple bioluminescent probe for in vitro detection of superoxide anion.
中文翻译：
使用水母发光蛋白生物发光检测超氧阴离子。
尽管超氧阴离子（O 2 –· ）是在正常细胞呼吸过程中产生的，并且在细胞增殖、迁移、凋亡和体内平衡等多种细胞过程中发挥重要作用，但其失调与多种疾病有关。鉴于这些在生物系统中的突出作用，超氧阴离子的准确定量方法的开发引起了极大的研究关注。在这里，我们评估了水母发光蛋白（一种钙依赖性发光蛋白）作为超氧阴离子的潜在生物发光报告蛋白。其机制基于水母发光蛋白生物发光的测量，超氧阴离子氧化下腔肠素浓度越低，水母发光蛋白再生量越低，导致生物发光减少。水母发光蛋白的生物发光强度与超氧阴离子浓度成正比，检测范围为4~40 000 pM，检测限（S/N = 3）为1.2 pM，比化学发光法低5000倍。该方法具有较高的灵敏度，已成功应用于植物细胞样品中超氧阴离子的测定。结果表明，基于发光蛋白的生物发光系统可以作为一种高度灵敏、特异且简单的生物发光探针，用于体外检测超氧阴离子。
更新日期：2019-10-02
中文翻译：
使用水母发光蛋白生物发光检测超氧阴离子。
尽管超氧阴离子（O 2 –· ）是在正常细胞呼吸过程中产生的，并且在细胞增殖、迁移、凋亡和体内平衡等多种细胞过程中发挥重要作用，但其失调与多种疾病有关。鉴于这些在生物系统中的突出作用，超氧阴离子的准确定量方法的开发引起了极大的研究关注。在这里，我们评估了水母发光蛋白（一种钙依赖性发光蛋白）作为超氧阴离子的潜在生物发光报告蛋白。其机制基于水母发光蛋白生物发光的测量，超氧阴离子氧化下腔肠素浓度越低，水母发光蛋白再生量越低，导致生物发光减少。水母发光蛋白的生物发光强度与超氧阴离子浓度成正比，检测范围为4~40 000 pM，检测限（S/N = 3）为1.2 pM，比化学发光法低5000倍。该方法具有较高的灵敏度，已成功应用于植物细胞样品中超氧阴离子的测定。结果表明，基于发光蛋白的生物发光系统可以作为一种高度灵敏、特异且简单的生物发光探针，用于体外检测超氧阴离子。
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