A research team led by Prof. MIAO Peng from the Suzhou Institute of Biomedical Engineering and Technology of the Chinese Academy of Sciences proposed a nano-impact electrochemical (NIE) sensing strategy for the highly sensitive determination of miRNA.
The new mechanism and results were published in Nano Letters entitled "Nano-Impact Electrochemical Biosensing Based on a CRISPR-Responsive DNA Hydrogel".
NIE enables the examination of the physicochemical properties and structure-activity relationships of nanomaterials at the nanometer scale. It contributes to the simple, rapid, and high-throughput analysis of biomolecules, but the relatively low effective collision frequency limits its broad prospects in basic research and clinical applications.
"We therefore proposed a novel NIE sensing strategy that was enhanced by the CRISPR-responsive DNA hydrogel and cascade DNA assembly, which significantly improved the sensitivity of this biosensor," said MIAO Peng.
The entire process included target miRNA-induced strand displacement amplification (SDA), catalytic hairpin assembly (CHA), and CRISPR/Cas trans-cutting.
By loading nanomaterials into DNA hydrogels, the limitation of one target triggering one collision entity in the traditional NIE mode could be overcome. In addition, the use of current frequency instead of peak current intensity mitigated the background interference and overcame the problem of poor stability in traditional electrochemical methods.
This biosensing strategy ensured ultra-high sensitivity, with a limit of detection for miR-141 as low as 4.21 aM. The researchers then measured the reactions triggered by four mismatched miRNAs and further tested the application of this method by challenging real samples. The method showed high selectivity and could successfully distinguish cells with abnormal miRNA levels.
This work has broad applicability in nucleic acid sensing and provides new insights into label-free and unmodified nano-electrochemical approaches, according to the researchers.
Fig. 1. (A) Illustration of target-induced SDA and CHA. (B) Illustration of DNA hydrogel formation and CRISPR/Cas-digested event for the electrochemical response of released AgNPs. (Image by SIBET)
Fig. 2. (A) Cyclic voltammograms of Au-SPE in 10 mM Na2S2O3 and 10 mM NaOH. (B) Cyclic voltammograms for each modification step. (C) Current–time curves of AgNPs for different SPEs. (D) Current–time experiments with and without AgNPs. (Image by SIBET)
Fig. 3. (A) Current–time curves for the detection of miR-141. (B) Linear relationship between the collision frequency and the logarithm of miR-141 concentration. (C) Selectivity of the NIE method. (D) NIE method for the detection of miRNA spiked in standard buffer, human serum, and cell conditions. (Image by SIBET)
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