Park Seonhwa, Singh Amardeep, Kim Sinyoung, Yang Haesik
Department of Chemistry and Chemistry Institute of Functional Materials, Pusan National University , Busan 609-735, Korea.
Anal Chem. 2014 Feb 4;86(3):1560-6. doi: 10.1021/ac403912d. Epub 2014 Jan 15.
We compare herein biosensing performance of two electroreduction-based electrochemical-enzymatic (EN) redox-cycling schemes [the redox cycling combined with simultaneous enzymatic amplification (one-enzyme scheme) and the redox cycling combined with preceding enzymatic amplification (two-enzyme scheme)]. To minimize unwanted side reactions in the two-enzyme scheme, β-galactosidase (Gal) and tyrosinase (Tyr) are selected as an enzyme label and a redox enzyme, respectively, and Tyr is selected as a redox enzyme label in the one-enzyme scheme. The signal amplification in the one-enzyme scheme consists of (i) enzymatic oxidation of catechol into o-benzoquinone by Tyr and (ii) electroreduction-based EN redox cycling of o-benzoquinone. The signal amplification in the two-enzyme scheme consists of (i) enzymatic conversion of phenyl β-d-galactopyranoside into phenol by Gal, (ii) enzymatic oxidation of phenol into catechol by Tyr, and (iii) electroreduction-based EN redox cycling of o-benzoquinone including further enzymatic oxidation of catechol to o-benzoquinone by Tyr. Graphene oxide-modified indium-tin oxide (GO/ITO) electrodes, simply prepared by immersing ITO electrodes in a GO-dispersed aqueous solution, are used to obtain better electrocatalytic activities toward o-benzoquinone reduction than bare ITO electrodes. The detection limits for mouse IgG, measured with GO/ITO electrodes, are lower than when measured with bare ITO electrodes. Importantly, the detection of mouse IgG using the two-enzyme scheme allows lower detection limits than that using the one-enzyme scheme, because the former gives higher signal levels at low target concentrations although the former gives lower signal levels at high concentrations. The detection limit for cancer antigen (CA) 15-3, a biomarker of breast cancer, measured using the two-enzyme scheme and GO/ITO electrodes is ca. 0.1 U/mL, indicating that the immunosensor is highly sensitive.
我们在此比较了两种基于电还原的电化学酶（EN）氧化还原循环方案[氧化还原循环与同时酶促扩增相结合（单酶方案）以及氧化还原循环与预先酶促扩增相结合（双酶方案）]的生物传感性能。为了使双酶方案中不必要的副反应最小化，分别选择β-半乳糖苷酶（Gal）和酪氨酸酶（Tyr）作为酶标记物和氧化还原酶，并且在单酶方案中选择Tyr作为氧化还原酶标记物。单酶方案中的信号放大包括：（i）Tyr将儿茶酚酶促氧化为邻苯醌，以及（ii）基于电还原的邻苯醌的EN氧化还原循环。双酶方案中的信号放大包括：（i）Gal将苯基β-D-吡喃半乳糖苷酶促转化为苯酚，（ii）Tyr将苯酚酶促氧化为儿茶酚，以及（iii）基于电还原的邻苯醌的EN氧化还原循环，包括Tyr将儿茶酚进一步酶促氧化为邻苯醌。通过将ITO电极浸入GO分散的水溶液中简单制备的氧化石墨烯修饰的铟锡氧化物（GO/ITO）电极，用于获得比裸ITO电极对邻苯醌还原更好的电催化活性。用GO/ITO电极测量的小鼠IgG的检测限低于用裸ITO电极测量时的检测限。重要的是，使用双酶方案检测小鼠IgG比使用单酶方案具有更低的检测限，因为尽管前者在高浓度时信号水平较低，但在低目标浓度时前者给出更高的信号水平。使用双酶方案和GO/ITO电极测量的乳腺癌生物标志物癌抗原（CA）15-3的检测限约为0.1 U/mL，表明该免疫传感器具有高灵敏度。
