Abstract:In this work, we constructed an exonuclease III cleavage reaction-based isothermal amplification of nucleic acids with CRISPR/Cas12a-mediated pH-induced regenerative Electrochemiluminescence (ECL) biosensor for ultrasensitive and specific detection of SARS-CoV-2 nucleic acids for SARS-CoV-2 diagnosis. The triple-stranded nucleic acid in this biosensor has an extreme dependence on pH, which makes our constructed biosensor reproducible. This is essential for effective large-scale screening of SARS-CoV-2 in areas… Show more
“…As previous literature reported [ 19 , 20 ], the UV–Vis spectrum of the g-C 3 N 4 /Au hybrid has a matching spectral overlap with the ECL emission spectrum of Ru centered at 460 nm, indicating that energy transfer from the g-C 3 N 4 /Au donor to the Ru acceptor can occur. Therefore, in the construction of the present sensor, only the emission peak of the ECL of g-C 3 N 4 (460 nm) needs to be excited, which excites the emission peak of Ru at 620 nm can be obtained using a filter.…”
In this work, we designed an ECL ratiometric biosensor with a three-stranded Y-type DNA (Y-DNA) probe and induced a hybridization chain reaction (HCR) for the highly sensitive detection of SARS-CoV-2 nucleic acid. The important component of this system is the self-assembled Y-Shaped probe based on three nucleic acids. Y1, Y2, and Y3 can be linked by complementary base pairing to Hairpin1 (H1), Hairpin2 (H2), and Ru modified DNA (Ru1), respectively. H1 and H2 can trigger the HCR reaction when activated by the SARS-CoV-2 RdRp gene and the 5′ end of Ru1. The 5′ end of Ru1 is modified with the Ru complex, which can produce a strong electrochemiluminescence luminescence signal at 620 nm under an applied voltage. Through the amplification of Y-DNA-induced HCR reaction, Ru1 on the electrode surface gradually increased, the ECL signal at 460 nm was gradually quenched, and the signal at 620 nm was steadily generated. The SARS-CoV-2 RdRp gene can be quantified according to the degree of decrease of ECL signal at 460 nm and the increase of ECL signal at 620 nm. Combining the two signal amplification strategies, this ratiometric ECL biosensor can accurately and efficiently detect the target gene with a detection limit of 59 aM.
“…As previous literature reported [ 19 , 20 ], the UV–Vis spectrum of the g-C 3 N 4 /Au hybrid has a matching spectral overlap with the ECL emission spectrum of Ru centered at 460 nm, indicating that energy transfer from the g-C 3 N 4 /Au donor to the Ru acceptor can occur. Therefore, in the construction of the present sensor, only the emission peak of the ECL of g-C 3 N 4 (460 nm) needs to be excited, which excites the emission peak of Ru at 620 nm can be obtained using a filter.…”
In this work, we designed an ECL ratiometric biosensor with a three-stranded Y-type DNA (Y-DNA) probe and induced a hybridization chain reaction (HCR) for the highly sensitive detection of SARS-CoV-2 nucleic acid. The important component of this system is the self-assembled Y-Shaped probe based on three nucleic acids. Y1, Y2, and Y3 can be linked by complementary base pairing to Hairpin1 (H1), Hairpin2 (H2), and Ru modified DNA (Ru1), respectively. H1 and H2 can trigger the HCR reaction when activated by the SARS-CoV-2 RdRp gene and the 5′ end of Ru1. The 5′ end of Ru1 is modified with the Ru complex, which can produce a strong electrochemiluminescence luminescence signal at 620 nm under an applied voltage. Through the amplification of Y-DNA-induced HCR reaction, Ru1 on the electrode surface gradually increased, the ECL signal at 460 nm was gradually quenched, and the signal at 620 nm was steadily generated. The SARS-CoV-2 RdRp gene can be quantified according to the degree of decrease of ECL signal at 460 nm and the increase of ECL signal at 620 nm. Combining the two signal amplification strategies, this ratiometric ECL biosensor can accurately and efficiently detect the target gene with a detection limit of 59 aM.
“…Scheme 1 illustrates the mechanism of the target-triggered cascade signal amplification for electrochemical assay of SARS-CoV-2 RNA. We have selected a 26-nucleotide sequence from the SARS-CoV-2 RdRp gene located in the ORF1ab region as the target [ 15 ], which can distinguish SARS-CoV-2 from other coronaviruses with high specificity [ 30 , 31 ]. The designed hairpin HP1 and HP2, which exist complementary sequences, can maintain independent stable structures separately, because their complementary sequences are blocked in their stems, respectively.…”
“…They reported a linear range of 10 aM–500 aM and a detection limit as low as 12.8 aM. Zhang et al [ 59 ] have reported the identification of the same gene (SARS-CoV-2 RNA-dependent RNA polymerase (RdRp)) using a pH engineered regenerative DNA tetrahedron and CRISPR-Cas12a as the amplification method ( Fig. 3 A).…”
Section: Ecl Biosensors For Viruses Detectionmentioning
confidence: 99%
“… Fig. 3 A: Schematic of the pH-induced biosensor for the detection of SARS-CoV-2 RdRp gene [ 59 ]. B: Scheme of the ECL DNA biosensor development [ 60 ].…”
Section: Ecl Biosensors For Viruses Detectionmentioning
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