Electrophoresis-Enhanced Detection of Deoxyribonucleic Acids on a Membrane-Based Lateral Flow Strip Using Avian Influenza H5 Genetic Sequence as the Model

Abstract

This study reports a simple strategy to detect a deoxyribonucleic acid (DNA) on a membrane-based lateral flow (MBLF) strip without tedious gel preparation, gel electrophoresis, and EtBr-staining processes. The method also enhances the detection signal of the genetic sample. A direct electric field was applied over two ends of the MBLF strips to induce an electrophoresis of DNAs through the strips. The signal enhancement was demonstrated by the detection of the H5 subtype of avian influenza virus (H5 AIV). This approach showed an excellent selectivity of H5 AIV from other two control species, Arabidopsis thaliana and human PSMA5. It also showed an effective signal repeatability and sensitivity over a series of analyte concentrations. Its detection limit could be enhanced, from 40 ng to 0.1 ng by applying 12 V. The nano-gold particles for the color development were labeled on the capture antibody, and UV-VIS and TEM were used to check if the labeling was successful. This detection strategy could be further developed to apply on the detection of drug-allergic genes at clinics or detection of infectious substances at incident sites by a simple manipulation with an aid of a mini-PCR machine and auxiliary kits.

Applications of gold nanoparticles in virus detection

“…Resonance light-scattering-based detections usually involve measuring the amount of light scattered by different light spectroscopy techniques, including localized surface plasmon resonance (LSPR) 52 , Raman spectroscopy 49 , 50 and dynamic light scattering (DLS) 51 , 53 . The colorimetric detection of viruses using AuNPs usually relies on two main techniques: 1) a color amplification technique in which AuNPs are applied to act as direct coloring labels with their characteristic, intense red color 55 , 58 , 59 , 66 ; and 2) color change technique in which a color change from red to purple occurs in response to particle aggregation. AuNP aggregation can be noncrosslinked aggregation caused by the target-triggered removal of stabilizing ligands from the surfaces of AuNPs 54 , 56 or interparticle-crosslinked aggregation caused by the binding of ligands on modified AuNPs with target analytes 57 , 67 .…”

“…The application of AuNPs in IAV detection has primarily been focused on developing assays that enable the direct detection of whole IAV particles 53 , 58 , 92 or viral RNA 64 , 66 , 67 , 70 , 71 , 83 , 86 through numerous AuNP-based fluorometric, SERS, scanometric, light-scattering, colorimetric, and electrochemical detection schemes (Table 1 ).…”

“…AuNP-based colorimetric assays have been developed for IAV detection through four main detection schemes: 1) The first scheme involves the application of AuNPs conjugated with anti-digoxigenin as colorimetric labels to detect the target viral DNA premodified with digoxigenin and electrophoresed through a membrane-based lateral-flow technique 66 . 2) The second scheme is simply based on the electrostatic interactions between the negatively charged phosphate groups of the target DNA amplified with LAMP and the positively charged CTAB bilayer on AuNRs, resulting in the aggregation of AuNRs and a color change from red to purple 67 .…”

Sensitivity enhancement in lateral flow assays: a systems perspective

“…Wu et al applied a direct electric field to selectively enhance the transport of nucleic acids, and improve sensitivity 400-fold, in an LFA that detected PCR products from an H5N1 influenza PCR assay. 19 Although the application of ICP to micro/nanofluidic platforms was recently reviewed, 20 there are some examples of ICP in paper-based devices. Gong et al demonstrated ICP in a saturated paper-based device, which improved sensitivity 5-fold for a FITC-BSA conjugate and 4-fold for bromocresol green dye compared to without ICP; while Liu et al demonstrated ICP in a Parafilm-and paper-based device, which concentrated FITC.…”

Rapid Detection of Gene HLA-A3101 on Membrane Based Lateral-Flow Strips

“…Our recent study revealed that an electric field could enhance the detection performance of single-trended Deoxyribonucleic acids (DNA) on the MBLF strips [20].…”