Ultrasensitive Multiplex Detection of Single Nucleotide Polymorphisms Based on Short-Chain Hybridization Combined with Online Preconcentration of Capillary Electrophoresis

Abstract

Reliable multiple single nucleotide polymorphisms (SNPs) detection at low abundance is of great significance for disease diagnosis and biomedical research. Herein, we have developed a novel and simple method for multiple SNPs detection combining solid-phase capture by specific hybridization with online preconcentration of capillary gel electrophoresis-laser-induced fluorescence (CGE-LIF). The method presents an excellent performance due to its favorable traits: the solid-phase short-chain hybridization ensures the high specificity of SNP detection; the effective separation ability of CGE can easily achieve multiplex detection; the simple online preconcentration significantly improves the detection sensitivity of fluorescent probe by nearly 100-fold. For a single SNP target, the assay achieves a limit of detection as low as 0.01–0.02% for three different NRAS mutations in the same codon. For multiple SNP targets, as low as 0.05% abundance can be easily realized. Our method is simple, efficient, ultrasensitive, and universal for multiple SNPs detection without complex enzymatic or chemical ligation reaction, which shows great potential in early clinical diagnosis.

Hairpin-Empowered Invasive Reaction Combined with Catalytic Hairpin Assembly Cascade Amplification for the Specific Detection of Single-Nucleotide Polymorphisms

“…The limit of detection (3 σ/slope) was estimated as 0.36 fM. As shown in Table S2, this double-amplified detection system based on hairpin-empowered invasive reaction and CHA cascade reaction had a distinct advantage in detection sensitivity compared with detection strategies reported in many literature. ,− …”

Capillary Electrophoresis: A Three-Year Literature Review

“…All of the PCR products were analyzed using CE with short-end injection, providing the advantages of high resolution and rapid separation. Zhou et al used solid-phase hybridization capture and CGE-LIF for the detection of multiple single nucleotide polymorphisms . FASI was used for the CGE online preconcentration, leading to a significant improvement in the detection sensitivity of the fluorescent probe by nearly 100-fold.…”

Recent Applications of Dynamic On-Line Sample Preconcentration Techniques in Capillary Electrophoresis

“…Since a high electric field is applied to the sample zone, the electrophoretic migration of ionic analytes in the sample zone is faster than that in the BGE, resulting in the stacking of the analytes to a narrow zone at the sample/BGS boundary. In recent years, FASS has been employed to the CE analysis of organic mercury [12], nitrate [13], catecholamines [14], amino acids [14,15], single nucleotide polymorphisms (SNPs) [16], β2-agonist bronchodilator [17], endogenous metabolites [18], imatinib [19], fluoxetine [20], and adamantane analogs [21] as summarized in Table 1. As a typical application, Zhou et al reported CGE analyses of SNPs.…”

“…As a typical application, Zhou et al reported CGE analyses of SNPs. By employing FASS, the detection sensitivity was improved to nearly 100-fold [16]. Wang et al combined FASS with on-line complexation using 3-mercapto-1-propanesulfonate for the speciation of organic mercury, which allowed the direct UV detection of three organic mercury complexes with the LOD of 11-16 ng/mL [12].…”