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Enhancing 3D DNA Walker-Induced CRISPR/Cas12a Technology for Highly Sensitive Detection of ExomicroRNA Associated with Osteoporosis

2024; American Chemical Society; Volume: 9; Issue: 3 Linguagem: Inglês

10.1021/acssensors.3c02533

2379-3694

Lijuan Luo, Fang Dong, Dandan Li, Xinmin Li, Xinyu Li, Yunpeng Fan, Caihong Qi, Jinyong Luo, Li Li, Bo Shen,

MicroRNA in disease regulation

Exosomal microRNAs (exomiRNAs) have emerged as promising biomarkers for the early clinical diagnosis of osteoporosis. However, their limited abundance and short length in peripheral blood present significant challenges for the accurate detection of exomiRNAs. Herein, we have designed and implemented an efficacious fluorescence-based biosensor for the highly sensitive detection of exomiRNA associated with osteoporosis, leveraging the enhancing 3D DNA walker-induced CRISPR/Cas12a technology. The engineered DNA walker is capable of efficiently transforming target exomiRNA into amplifying DNA strands, thereby enhancing the sensitivity of the developed biosensor. Concurrently, the liberated DNA strands serve as activators to trigger Cas12a trans-cleavage activity, culminating in a significantly amplified fluorescent signal for the highly sensitive detection of exomiRNA-214. Under optimal conditions, the devised technology demonstrated the capacity to detect target exomiRNA-214 at concentrations as low as 20.42 fM, encompassing a wide linear range extending from 50.0 fM to 10.0 nM. Moreover, the fluorescence-based biosensor could accurately differentiate between healthy individuals and osteoporosis patients via the detection of exomiRNA-214, which was in agreement with RT-qPCR results. As such, this biosensing technology offers promise as a valuable tool for the early diagnosis of osteoporosis.