Pasquardini L, Potrich C, Vaghi V, Lunelli L, Frascella F, Descrovi E, Pirri C F, Pederzolli C
FBK - Fondazione Bruno Kessler, Laboratory of Biomolecular Sequence and Structure Analysis for Health, via Sommarive 18, I-38123 Povo (Trento), Italy.
FBK - Fondazione Bruno Kessler, Laboratory of Biomolecular Sequence and Structure Analysis for Health, via Sommarive 18, I-38123 Povo (Trento), Italy; CNR - Consiglio Nazionale delle Ricerche, Istituto di Biofisica, via alla Cascata 56/C, I-38123 Povo (Trento), Italy.
Talanta. 2016 Apr 1;150:699-704. doi: 10.1016/j.talanta.2016.01.002. Epub 2016 Jan 16.
The detection of low abundant biomarkers, such as circulating microRNAs, demands innovative detection methods with increased resolution, sensitivity and specificity. Here, a biofunctional surface was implemented for the selective capture of microRNAs, which were detected through fluorescence enhancement directly on a photonic crystal. To set up the optimal biofunctional surface, epoxy-coated commercially available microscope slides were spotted with specific anti-microRNA probes. The optimal concentration of probe as well as of passivating agent were selected and employed for titrating the microRNA hybridization. Cross-hybridization of different microRNAs was also tested, resulting negligible. Once optimized, the protocol was adapted to the photonic crystal surface, where fluorescent synthetic miR-16 was hybridized and imaged with a dedicated equipment. The photonic crystal consists of a dielectric multilayer patterned with a grating structure. In this way, it is possible to take advantage from both a resonant excitation of fluorophores and an angularly redirection of the emitted radiation. As a result, a significant fluorescence enhancement due to the resonant structure is collected from the patterned photonic crystal with respect to the outer non-structured surface. The dedicated read-out system is compact and based on a wide-field imaging detection, with little or no optical alignment issues, which makes this approach particularly interesting for further development such as for example in microarray-type bioassays.
低丰度生物标志物（如循环微小核糖核酸）的检测需要创新的检测方法，以提高分辨率、灵敏度和特异性。在此，实现了一种生物功能表面用于选择性捕获微小核糖核酸，这些微小核糖核酸通过直接在光子晶体上的荧光增强来检测。为了建立最佳的生物功能表面，用特定的抗微小核糖核酸探针点样涂有环氧树脂的市售显微镜载玻片。选择并使用探针以及钝化剂的最佳浓度来滴定微小核糖核酸杂交。还测试了不同微小核糖核酸的交叉杂交，结果可忽略不计。一旦优化，该方案就适用于光子晶体表面，荧光合成的miR-16在该表面杂交并用专用设备成像。光子晶体由具有光栅结构的介质多层构成。通过这种方式，可以利用荧光团的共振激发和发射辐射的角度重定向。结果，相对于外部非结构化表面，从图案化的光子晶体收集到由于共振结构而产生的显著荧光增强。该专用读出系统紧凑，基于宽场成像检测，几乎没有光学对准问题，这使得这种方法对于进一步发展（例如在微阵列型生物测定中）特别有吸引力。
