Beijing National Laboratory for Molecular Sciences, Key Laboratory of Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese Academy of Sciences , Beijing 100190, China.
University of the Chinese Academy of Sciences , Beijing 100049, China.
ACS Appl Mater Interfaces. 2017 Mar 22;9(11):9462-9469. doi: 10.1021/acsami.7b00418. Epub 2017 Mar 10.
We present a novel microbead-based detection platform as a simple and universal strategy for simultaneous determination of multiple biomolecules. This platform is composed of streptavidin coated uniform-sized polystyrene microbeads, dye and biotin-labeled ssDNA or aptamer probes, and quencher-labeled complementary sequences. By this method, upon target binding to the probes, quencher strand dissociation is triggered, which results in fluorescence reactivation of the microbead linked probes. The fluorescence variation is readily monitored by flow cytometry and with a high sensitivity. Explicitly, this microbead-based detection platform shows a high sensitivity for target DNA with a detection limit as low as 0.20 nM, alongside good selectivity from one-base mismatched DNA. This novel platform also shows good selectivity and high sensitivity for protein detection when aptamer is used as a probe. The detection limit for lysozyme is as low as 8.56 nM. Moreover, simultaneous detection of multiple targets has been achieved via incorporating different dye-labeled probes on the microbeads concurrently. We have also applied this developed strategy to the detection of target DNA in human serum. This strategy can be easily extended to other targets through simple probe and quencher variation.
我们提出了一种基于微球的新型检测平台，作为一种简单而通用的策略，用于同时测定多种生物分子。该平台由链霉亲和素包被的均一尺寸聚苯乙烯微球、染料和生物素标记的 ssDNA 或适体探针、以及淬灭剂标记的互补序列组成。通过这种方法，在目标物与探针结合后，引发淬灭剂链解离，导致与微球连接的探针的荧光重新激活。通过流式细胞术很容易监测到荧光变化，并具有很高的灵敏度。具体而言，这种基于微球的检测平台对目标 DNA 具有很高的灵敏度，检测限低至 0.20 nM，并且对单碱基错配 DNA 具有良好的选择性。当适体用作探针时，该新型平台在蛋白质检测中也表现出良好的选择性和高灵敏度。溶菌酶的检测限低至 8.56 nM。此外，通过同时将不同染料标记的探针结合到微球上，实现了对多个目标物的同时检测。我们还将这种开发的策略应用于人血清中目标 DNA 的检测。通过简单地改变探针和淬灭剂，这种策略可以很容易地扩展到其他目标物。
