Jingwen Zhu,Xia Zong,Taijing Xu,Ying Chen,Xinjue Dai,Xiaowei Cao,Changhua Lu
Early detection of biomarkers has a crucial role in preventing blindness caused by diabetic retinopathy (DR). In this work, we developed a rapid, highly sensitive method for the simultaneous detection of DR-related biomarkers (miR-21 and miR-152) based on an Au nano-hexagon (AuNHs) integrated microarray chip combined with a competitive recognition strategy. The nucleic acid aptamer-modified AuNHs array served as capture substrate, while Au nano-trisoctahedrons (AuNTs) with complementary strands of nucleic acid aptamers mounted on their surface and Raman signaling molecules served as SERS probes (AuNTs@4-MBA@H1 and AuNTs@DTNB@H2). SERS probes carrying Raman signaling molecules (4-MBA and DTNB) were displaced by the target miRNAs in the experiments because they had a higher binding affinity to nucleic acid aptamers than their structural analogs (H1 and H2). This approach led to a notable decrease in the Raman signal. The competitive recognition strategy was straightforward and useful, and required only a one-step dropwise reaction to detect both miRNAs simultaneously and effectively. The microarray chip designed accordingly had the advantages of sensitive, rapid and high-throughput detection, and the detection time was only 8 min. The limits of detection (LOD) for miR-21 and miR-152 were as low as 2.76 × 10−14 M and 1.94 × 10−14 M, respectively. The utility of the method in complex systems was verified by detecting miRNAs in the serum of DR rats, and the accuracy of the method was verified against the detection results of a RT-qPCR method. This method provides a new idea and manner for the detection of miRNAs in serum, which is crucial for the early diagnosis of DR.
Shuang Tan,Rui Han,Shilian Wu,Huan Liang,Yuting Zhao,Hui Zhao,Can-Peng Li
Competitive host-guest recognition has been utilized to determine small molecules using macrocyclic supramolecular host, while less studies focused on the specific recognition and sensing of protein. In the present work, we are the first time to report a label-free fluorescent assay for insulin determination based on the supramolecular recognition between cationic pillar[6]arene (CP6) and insulin. The approach is based on fluorescence resonance energy transfer (FRET) through competitive recognition between CP6 functionalized reduced graphene oxide (CP6@rGO) and probe/insulin molecules. Probe molecule (RhB) has strong fluorescent signal, and its fluorescent is quenched by rGO based on FRET. When target protein molecule (insulin) is added to CP6@rGO, the probe is displaced by insulin and a host-guest complex CP6@rGO/insulin is formed, resulting in a “turn-on” fluorescence signal. The fluorescence intensity of complex increased linearly with the increase of insulin concentration ranging 0.01–0.50 and 1.0–16.0 μM, respectively with a detection limit of 3 nM. The sensor was successfully utilized to determine insulin in artificial serum. The molecular docking result showed that the N-terminal Phe of insulin's B chain was included in the CP6 cavity through electrostatic interaction and formed a stable host-guest complex.
Zhiwei Qie,Qiqi Liu,Wenliang Yan,Zichen Gao,Wu Meng,Rui Xiao,Shengqi Wang
A universal and ultrasensitive immunochromatographic assay (ICA) was established using antigen as a bifunctional element and antialbumin antibody in a test line. Preincubation was introduced for competitive recognition. After optimization, the linear detection of aflatoxin M1 (AFM1) with quantum dot bead (QB)-based ICA (QB-ICA) sensor ranged from 10 to 52 pg mL–1, with a 50% inhibitory concentration (IC50) of 23 pg mL–1, which was nearly 49.6-fold lower than those of ICA on a traditional structure with traditional pretreatment (IC50 = 1.10 ng mL–1) and 10-fold lower than those of ICA on a traditional structure with acid aid pretreatment (IC50 = 0.25 ng mL–1). The limit of detection (LOD) for AFM1 was 16 pg mL–1 in milk, which was approximately 16.3-fold times higher than those of ICA on a traditional structure with traditional pretreatment and 6.3-fold higher than those of ICA on a traditional structure with acid aid pretreatment. The LOD improved by 20-fold by using the proposed structure compared to that of conventional enzyme-linked immunosorbent assay (ELISA) for AFM1-spiked milk samples (IC10 = 0.12 ng mL–1). The performance and practicability of the established QB-ICA sensor were validated with a commercial ELISA kit. To evaluate universality, we successfully detected chloramphenicol, with IC50 of 0.42 ng mL–1. Given its high sensitivity and universality, the proposed QB-ICA can be used as an alternative for rapid, sensitive, and universal quantitative detection of all small-molecule analytes.
危险品化学品经营许可证（带存储）