Browsing by Author "Zhang, Pengbo"

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Binding Studies of Cationic Conjugated Polymers and DNA for Label-Free Fluorescent Biosensors
(American Chemical Society, 2022-07-19) Zhang, Pengbo; Lu, Chang; Niu, Chenqi; Wang, Xiaoyu; Li, Zhengping; Liu, Juewen
Cationic conjugated polymers (CCPs), especially polythiophene, have been extensively used as probes for developing DNA and aptamer-based biosensors. Although many interesting applications have been achieved, a fundamental understanding of this system remains quite limited. In this work, we performed systematic binding assays to understand the interactions between poly(3-(3′-N,N,N-triethylamino-1′-propyloxy)-4-methyl-2,5-thiophene) (PMNT) and DNA. The fluorescence of PMNT at 530 nm initially decreased and then a peak at 580 nm emerged after binding with single-stranded DNA (ssDNA). The binding force between PMNT and DNA was dominated by electrostatic interactions at first and then DNA base-mediated interactions also became important. Since the bases in double-stranded DNA (dsDNA) were shielded, their fluorescence changes were quite different. To best differentiate ssDNA and dsDNA, the optimal pH was between 6 and 8, and the optimal NaCl concentration was around 0.3 M. Moreover, by changing the sequence and length of ssDNA, poly-T had the largest fluorescence shift and poly-A had the smallest change. Under the optimized conditions, the PMNT-based biosensor had a detection limit of 1 nM DNA, which was similar to the SYBR Green I-based assay.
General Label-free Fluorescent Aptamer Binding Assay Using Cationic Conjugated Polymers
(American Chemical Society, 2022-10-25) Zhang, Pengbo; Qin, Ke; Lopez, Anand; Li, Zhengping; Liu, Juewen
With more and more new aptamers being reported, a general, cost-effective yet reliable aptamer binding assay is still needed. Herein, we studied cationic conjugated polymer (CCP)-based binding assays taking advantage of the conformational change of aptamer after binding with a target, which is reflected by the fluorescence change of the CCP. Poly(3-(3′-N,N,N-triethylamino-1′-propyloxy)-4-methyl-2,5-thiophene hydrochloride) (PMNT) was used as a model CCP in this study, and the optimal buffer was close to physiological conditions with 100 mM NaCl and 10 mM MgCl2. We characterized four aptamers for K+, adenosine, cortisol, and caffeine. For cortisol and caffeine, the drop in the 580 nm peak intensity was used for quantification, whereas for K+ and adenosine, the fluorescence ratio at 580 over 530 nm was used. The longer stem of the stem-loop structured aptamer facilitated binding of the target and enlarged the detection signal. High specificity was achieved in differentiating targets with analogues. Compared with the SYBR Green I dye-based staining method, our method achieved equal or even higher sensitivity. Therefore, this assay is practicable as a general aptamer binding assay. The simple, label-free, quick response, and cost-effective features will make it a useful method to evaluate aptamer binding. At the same time, this system can also serve as label-free biosensors for target detection.