Mirasoli Mara, Nascetti Augusto, Caputo Domenico, Zangheri Martina, Scipinotti Riccardo, Cevenini Luca, de Cesare Giampiero, Roda Aldo
Department of Chemistry "G. Ciamician,", University of Bologna, 40126, Bologna, Italy,
Anal Bioanal Chem. 2014 Sep;406(23):5645-56. doi: 10.1007/s00216-014-7971-9. Epub 2014 Jun 26.
We propose a disposable multiwell microcartridge with integrated amorphous silicon photosensors array for bio- and chemiluminescence-based bioassays, where the enzymatic reactions and the detection unit are coupled on the same glass substrate. Each well, made in a polydimethylsiloxane (PDMS) unit, hosts an enzymatic reaction that is monitored by one photosensor of the array. Photosensors were characterized in terms of their dark current background noise and response to different wavelengths of visible light in order to determine their suitability as detection devices for chemical luminescent phenomena. Calibration curves of the photosensors' response to different luminescent systems were then evaluated by using the chemiluminescent reactions catalyzed by alkaline phosphatase and horseradish peroxidase and the bioluminescent reaction catalyzed by firefly luciferase. Limits of detection in the order of attomoles for chemiluminescence enzymes and femtomoles for luciferase and sensitivities in the range between 0.007 and 0.1 pA pmol(-1) L were reached. We found that, without the need of cooling systems, the analytical performances of the proposed cartridge are comparable with those achievable with state-of-the-art thermoelectrically cooled charge-coupled device-based laboratory instrumentation. In addition, thanks to the small amount of generated output data, the proposed device allows the monitoring of long-lasting reactions with significant advantages in terms of data-storage needs, transmission bandwidth, ease of real-time signal processing and limited power consumption. Based on these results, the operation in model bioanalytical assays exploiting luminescent reactions was tested demonstrating that a-Si:H photosensors arrays, when integrated with PDMS microfluidic units, provide compact, sensitive and potentially low-cost microdevices for chemiluminescence and bioluminescence-based bioassays with a wide range of possible applications for in-field and point-of-care bio-analyses.
我们提出了一种带有集成非晶硅光电传感器阵列的一次性多孔微盒，用于基于生物发光和化学发光的生物检测，其中酶促反应和检测单元耦合在同一玻璃基板上。每个孔由聚二甲基硅氧烷（PDMS）单元制成，容纳一个酶促反应，该反应由阵列中的一个光电传感器进行监测。对光电传感器的暗电流背景噪声以及对不同可见光波长的响应进行了表征，以确定它们作为化学发光现象检测设备的适用性。然后，通过使用碱性磷酸酶和辣根过氧化物酶催化的化学发光反应以及萤火虫荧光素酶催化的生物发光反应，评估了光电传感器对不同发光系统响应的校准曲线。化学发光酶的检测限达到阿托摩尔级别，荧光素酶的检测限达到飞摩尔级别，灵敏度在0.007至0.1 pA pmol⁻¹ L范围内。我们发现，无需冷却系统，所提出的微盒的分析性能与基于最先进的热电冷却电荷耦合器件的实验室仪器所实现的性能相当。此外，由于产生的输出数据量少，所提出的设备能够监测持久反应，在数据存储需求、传输带宽、实时信号处理的简易性和低功耗方面具有显著优势。基于这些结果，测试了利用发光反应的模型生物分析测定中的操作，证明当与PDMS微流体单元集成时，非晶硅氢化（a-Si:H）光电传感器阵列可为基于化学发光和生物发光的生物检测提供紧凑、灵敏且潜在低成本的微型设备，在现场和即时护理生物分析中有广泛的可能应用。
