Development of fluorescent nanoparticle-labeled lateral flow assay for the detection of nucleic acids

Abstract

The rapid, specific and sensitive detection of nucleic acids is of utmost importance for the identification of infectious agents, diagnosis and treatment of genetic diseases, and the detection of pathogens related to human health and safety. Here we report the development of a simple and sensitive nucleic acid sequence-based and Ru(bpy)3 (2+)-doped silica nanoparticle-labeled lateral flow assay which achieves low limit of detection by using fluorescencent nanoparticles. The detection of the synthetic nucleic acid sequences representative of Trypanosoma mRNA, the causative agent for African sleeping sickness, was utilized to demonstrate this assay. The 30 nm spherical Ru(bpy)3 (2+)-doped silica nanoparticles were prepared in aqueous medium by a novel method recently reported. The nanoparticles were modified by 3-glycidoxypropyl trimethoxysilane in order to conjugate to amine-capped oligonucleotide reporter probes. The fluorescent intensities of the fluorescent assays were quantified on a mictrotiter plate reader using a custom holder. The experimental results showed that the lateral flow fluorescent assay developed was more sensitive compared with the traditional colloidal gold test strips. The limit of detection for the fluorescent lateral flow assay developed is approximately 0.066 fmols as compared to approximately 15 fmols for the colloidal gold. The limit of detection can further be reduced about one order of magnitude when "dipstick" format was used.

Nanoparticle-based Point of Care Immunoassays for in vitro Biomedical Diagnostics

“…27,28 AuNPs have been used to detect and screen many analytes, including DNA, protein, enzymes, hormones, ions and cancer biomarkers. [29][30][31][32][33][34] The distinct optical properties of AuNPs have been coupled with the catalytic amplification of the traditional enzymatic chromogenic agent horseradish peroxidase (HRP) to develop a dual label (i.e. HRP-AuNP) assay for lateral flow nucleic acid biosensors (LFNAB) for hybridization-based ultrasensitive naked eye detection of DNA (Figs.…”

Recent Advances in Nanoparticles-based Lateral Flow Biosensors

“…In addition, the use of fluorescent technique [98][99][100][101][102][103][104][105], up-converting phosphorus [106,107,59,74] and technology also allowed an increase in sensitivity and simplicity compared with that for visual detection of traditional colored labels.…”

Fluorescent carbon nanoparticle-based lateral flow biosensor for ultrasensitive detection of DNA

“…Fluorescent nanoparticles are a great alternative to counterfeit the problems of sensitivity associated with colorimetric detections (Zheng et al, 2012 and Kim et al, 2011), Quantum dots (Li et al, 2010), fluorophore-labeled sequences (Xu et al, 2014), dye-doped silica nanoparticles (Wang and Nugen., 2013) and up-converting phosphor reporters (Corstjens et al, 2003) have been used as fluorescent tags to prepare the fluorescent lateral flow biosensors for ultrasensitive detection of nucleic acid sequences. The sensitivities of the fluorescent lateral flow nucleic acid biosensors without additional signal amplification were 1–3 orders higher than that of GNP-based colorimetric lateral flow biosensors.…”