Enzyme-Free Colorimetric Detection of DNA by Using Gold Nanoparticles and Hybridization Chain Reaction Amplification

Abstract

A novel, high sensitive, and specific DNA assay based on gold nanoparticle (AuNP) colorimetric detection and hybridization chain reaction (HCR) amplification has been demonstrated in this article. Two hairpin auxiliary probes were designed with single-stranded DNA (ssDNA) sticky ends which stabilize AuNPs and effectively prevent them from salt-induced aggregation. The target DNA hybridized with the hairpin auxiliary probes and triggered the formation of extended double-stranded DNA (dsDNA) polymers through HCR. As a result, the formed dsDNA polymers provide less stabilization without ssDNA sticky ends, and AuNPs undergo aggregation when salt concentration is increased. Subsequently, a pale purple-to-blue color variation is observed in the colloid solution. The system is simple in design and convenient in operation. The novel strategy eliminates the need for enzymatic reactions, separation processes, chemical modifications, and sophisticated instrumentation. The detection and discrimination process can be seen with the naked eye. The detection limit of this method is lower than or at least comparable to previous AuNP-based methods. Importantly, the protocol offers high selectivity for the determination between perfectly matched target oligonucleotides and targets with single base-pair mismatches.

Novel Signal-Enhancing Approaches for Optical Detection of Nucleic Acids—Going beyond Target Amplification

“…In a recent paper, Au-nps were incorporated into a hybridization chain reaction assay for SNP detection in model DNA targets. Remarkably, they achieved a LOD of 50 pM with instrument detection and 100 pM with naked eye observation, which is about two orders of magnitude lower than that of previously reported Au-np-based assays [50].…”

“…1 kg in weight) and is compatible with most labels and aforementioned signal-enhancing techniques. Using a simple photoresistor as a signal detector the LOD is 50 pM of DNA [50]. However, colorimetry can only be applied when the probe's absorbance wavelengths lie within the visible region [57].…”

“…PMTs contain a photocathode, several dynodes, and an anode. [64] Xe = xenon; MH = metal halide; (HI) LED = (high intensity) light emitting diode; PR = photoresisitor; PD = photodiode; EPC = electronic photon counter; CCD, CID and CMOS = charge-coupled device, charge-injection device and complementary-metal-oxide-semiconductor device; EBCCD = electron-bombarded CCD; GaAsP = Gallium Arsenide Phosphide); FLS and LSC = fluorescence and luminescence; QY = quantum yield; * LOD values are reported for particular assays described in the cited literature [50,[57][58][59][60][61][62]64]. For RT-qPCR, LOD is given for amplified DNA product.…”

Sensitive Colorimetric Detection of MicroRNA Based on Target Catalyzed Double-arm Hairpin DNA Assembling

“…The linear-regression equation was A630/A520 = 0.1828 log c + 0.8696 (R = 0.9901), and the detection limit that was taken to be three-times the standard derivation in a blank solution is 3 pM, which is lower than that of the colorimetric detection of DNA previously reported. 33 This may be because the double-arm hairpin probe DNA could protect AuNP from aggregation more efficiently, and thus reduce the background signal, besides, that the length of the sticky ends of the obtained double-stranded DNA may also influence the detection sensitivity.…”

A new colorimetric platform for ultrasensitive detection of protein and cancer cells based on the assembly of nucleic acids and proteins

“…The initiator can trigger a cascade of hybridization events to yield nicked double helices analogous to alternating copolymers, [20,21] giving HCR great potential for signal amplification [22][23][24]. More recently, some researchers have combined the amplification capability of HCR with colorimetric methods for the detection of proteins [25,26] due to the simplicity and the lack of requiring expensive instruments.…”