3D DNA Walker-Assisted CRISPR/Cas12a Trans-Cleavage for Ultrasensitive Electrochemiluminescence Detection of miRNA-141

Abstract

In this study, a CRISPR/Cas12a (LbCpf1)mediated electrochemiluminescence (ECL) paper-based platform on the basis of a three-dimensional (3D) DNA walker was proposed for the ultrasensitive detection of miRNA-141. Initially, 3D-rGO with a tremendous loading space was modified on the paper working electrode (PWE) to construct an excellent conductive substrate and facilitate the growth of AuPd nanoparticles (NPs). Afterward, the AuPd NPs were introduced as the coreaction emitter medium of the 3D-rGO/PWE to provide convenience for the transformation between S 2 O 8 2− and SO 4 2− , amplifying the ECL emission of g-C 3 N 4 nanosheets (NSs). Meanwhile, with the help of Nt.BsmAI nicking endonuclease, a 3D DNA walker signal amplifier was designed to convert and magnify the target miRNA-141 into a particular trigger sequence, which could act as activator DNA to motivate the trans-acting deoxyribonuclease activity of CRISPR/Cas12a to further achieve efficient annihilation of the ECL signal. Furthermore, the proposed multimechanism-driven biosensor exhibited excellent sensitivity and specificity, with a relatively low detection limit at 0.331 fM (S/ N = 3) in the concentration range between 1 fM and 10 nM. Consequently, the designed strategy not only extended the application scope of CRISPR/Cas12a but also devoted a new approach for the clinical diagnosis of modern medicine.

Ultrasensitive detection of exosomal miRNA with PMO-graphene quantum dots-functionalized field-effect transistor biosensor

“…In order to better show the sensitivity of the sensor, we compared detection range and LOD of the GPPR-FET sensor with those of sensors ( Caputo et al., 2019 ; Cogal et al., 2021 ; Gao et al., 2020 ; Huang et al., 2021 ; Kim et al., 2021 ; Li et al., 2021 ; Wang et al., 2021 ; Wu et al., 2021 ). As shown in Table S2 , it can be seen that the method established in this article is superior to other sensors in terms of detection range and LOD.…”

Next-Generation Diagnostic with CRISPR/Cas: Beyond Nucleic Acid Detection

“…Besides Cas9, several CRISPR/Cas13 and Cas12-based COVID-19 detection platform such as STOPCovid, CREST, SHINE, SHERLOCK and DETECTR for COVID, VanGuard, AIOD, DISCoVER and many more have been developed for the same purpose [ 1 , 62 , 63 , 64 , 65 , 75 , 77 , 103 , 104 ]. Various CRISPR/Cas-based platforms aided with isothermal amplification have been utilized for the detection of Porcine reproductive and respiratory syndrome virus (PRRSV) [ 67 ], ASFV [ 32 , 105 ], Zika/Dengue virus [ 73 ], HIV [ 65 ], HPV16/18 [ 66 , 96 ] BK polyomavirus and Cytomegalovirus [ 106 ].…”

Peroxidase-Like N-Doped Carbon Nanotubes Loaded with NiCo-Layered Double-Hydroxide Nanoflowers for Dual-Mode Ultrasensitive and Real-Time Detection of H2O2 Secreted from Cancer Cells

“…Therefore, a dual-mode signal strategy is required to avoid false positives for single signals. In addition, the sensitivity of electrochemiluminescence (ECL) is much higher than that of the electrochemical method. − H 2 O 2 can significantly enhance the intensity of luminol at extremely low levels, expanding the use of ECL for ultra-sensitive detection and imaging of H 2 O 2 . Moreover, imaging detection of H 2 O 2 , glucose, and cholesterol in a single cell has been achieved using luminol and its derivatives, L-012. − Recently, simultaneous electrochemical and ECL analyses have been reported.…”