Extensive research is focused on the development of highly sensitive, rapid on-site diagnostic devices. The lateral flow strip (LFS) is a paper-based point-of-care diagnostic device, which is highly promising because of its ease of use and low cost. Despite these advantages, LFS device is still less popular than other methods such as enzyme-linked immunosorbent assay (ELISA) or real-time polymerase chain reaction (qPCR) due to its low sensitivity. Here, we have developed a fluorescence-based lateral flow strip (f-LFS) device for DNA detection using a molecular beacon (MB), a short hairpin-forming DNA strand tagged with a fluorophore-quencher pair. Each paper and membrane component of f-LFS device was carefully selected based on their physicochemical properties including porosity, surface functionality, and autofluorescence. The limit of detection (LOD) of this device was substantially improved to 2.1 fg/mL by adding MgCl 2 to the reaction buffer and narrowing the test membrane dimension. Also, a portable fluorescence detection system for f-LFS was developed using a multi-pixel photon counter (MPPC), a sensitive detector detecting the signal on site. We anticipate that this highly sensitive paper-based diagnostic device can be utilized for on-site diagnosis of various diseases.
more »
« less
Label-free detection of DNA hybridization with a compact LSPR-based fiber-optic sensor
A miniaturized, robust, localized surface plasmon resonance (LSPR)-coupled fiber-optic (FO) nanoprobe providing an integrated and portable solution for detection of DNA hybridization and measurement of DNA concentrations has been demonstrated. The FO nanoprobe was created by constructing arrays of metallic nanostructures on the end facets of optical fibers utilizing nanofabrication technologies, including electron beam lithography and lift-off processes. The LSPR-FO nanoprobe device offers real-time, label-free, and low-sample-volume quantification of single-strand DNA in water with high sensitivity and selectivity, achieving a limit of detection around 10 fM. These results demonstrate the feasibility of the LSPR-FO nanoprobe device as a compact and low-cost biosensor for detection of short-strand DNA.
more »
« less
- Award ID(s):
- 1511194
- PAR ID:
- 10111221
- Date Published:
- Journal Name:
- The Analyst
- Volume:
- 142
- Issue:
- 11
- ISSN:
- 0003-2654
- Page Range / eLocation ID:
- 1974 to 1981
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
This paper proposes an iterative method of estimating power system forced oscillation (FO) amplitude, frequency, phase, and start/stop times from measured data. It combines three algorithms with favorable asymptotic statistical properties: a periodogram-based iterative frequency estimator, a Discrete-Time Fourier Transform (DTFT)-based method of estimating amplitude and phase, and a changepoint detection (CPD) method for estimating the FO start and stop samples. Each of these have been shown in the literature to be approximate maximum likelihood estimators (MLE), meaning that for large enough sample size or signal-to-noise ratio (SNR), they can be unbiased and reach the Cramer-Rao Lower Bound in variance. The proposed method is shown through Monte Carlo simulations of a low-order model of the Western Electricity Coordinating Council (WECC) power system to achieve statistical efficiency for low SNR values. The proposed method is validated with data measured from the January 11, 2019 US Eastern Interconnection (EI) FO event. It is shown to accurately extract the FO parameters and remove electromechanical mode meter bias, even with a time-varying FO amplitude.more » « less
-
Transcription factor (TF) target search on genome is highly essential for gene expression and regulation. High-resolution determination of TF diffusion along DNA remains technically challenging. Here, we constructed a TF model system using the plant WRKY domain protein in complex with DNA from crystallography and demonstrated microsecond diffusion dynamics of WRKY on DNA by employing all-atom molecular-dynamics (MD) simulations. Notably, we found that WRKY preferentially binds to one strand of DNA with significant energetic bias compared with the other, or nonpreferred strand. The preferential DNA-strand binding becomes most prominent in the static process, from nonspecific to specific DNA binding, but less distinct during diffusive movements of the domain protein on the DNA. Remarkably, without employing acceleration forces or bias, we captured a complete one-base-pair stepping cycle of the protein tracking along major groove of DNA with a homogeneous poly-adenosine sequence, as individual hydrogen bonds break and reform at the protein–DNA binding interface. Further DNA-groove tracking motions of the protein forward or backward, with occasional sliding as well as strand crossing to minor groove of DNA, were also captured. The processive diffusion of WRKY along DNA has been further sampled via coarse-grained MD simulations. The study thus provides structural dynamics details on diffusion of a small TF domain protein, suggests how the protein approaches a specific recognition site on DNA, and supports further high-precision experimental detection. The stochastic movements revealed in the TF diffusion also provide general clues about how other protein walkers step and slide along DNA.more » « less
-
Abstract DNA has emerged as a powerful substrate for programming information processing machines at the nanoscale. Among the DNA computing primitives used today, DNA strand displacement (DSD) is arguably the most popular, with DSD-based circuit applications ranging from disease diagnostics to molecular artificial neural networks. The outputs of DSD circuits are generally read using fluorescence spectroscopy. However, due to the spectral overlap of typical small-molecule fluorescent reporters, the number of unique outputs that can be detected in parallel is limited, requiring complex optical setups or spatial isolation of reactions to make output bandwidths scalable. Here, we present a multiplexable sequencing-free readout method that enables real-time, kinetic measurement of DSD circuit activity through highly parallel, direct detection of barcoded output strands using nanopore sensor array technology (Oxford Nanopore Technologies’ MinION device). These results increase DSD output bandwidth by an order of magnitude over what is currently feasible with fluorescence spectroscopy.more » « less
-
Abstract Watson–Crick base pairing rules provide a powerful approach for engineering DNA‐based nanodevices with programmable and predictable behaviors. In particular, DNA strand displacement reactions have enabled the development of an impressive repertoire of molecular devices with complex functionalities. By relying on DNA to function, dynamic strand displacement devices represent powerful tools for the interrogation and manipulation of biological systems. Yet, implementation in living systems has been a slow process due to several persistent challenges, including nuclease degradation. To circumvent these issues, researchers are increasingly turning to chemically modified nucleotides as a means to increase device performance and reliability within harsh biological environments. In this review, we summarize recent progress toward the integration of chemically modified nucleotides with DNA strand displacement reactions, highlighting key successes in the development of robust systems and devices that operate in living cells and in vivo. We discuss the advantages and disadvantages of commonly employed modifications as they pertain to DNA strand displacement, as well as considerations that must be taken into account when applying modified oligonucleotide to living cells. Finally, we explore how chemically modified nucleotides fit into the broader goal of bringing dynamic DNA nanotechnology into the cell, and the challenges that remain. This article is categorized under:Diagnostic Tools > In Vivo Nanodiagnostics and ImagingNanotechnology Approaches to Biology > Nanoscale Systems in BiologyDiagnostic Tools > Biosensingmore » « less
An official website of the United States government

