Laboratory of Bioanalysis, Institute of Biochemistry & Biophysics, University of Tehran, Tehran, Iran.
Gastroenterology and Liver Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
Mikrochim Acta. 2024 Oct 3;191(11):644. doi: 10.1007/s00604-024-06688-4.
A low-cost, lab-made polytetrafluoroethylene micro-cell, equipped with three electrodes, wasd eveloped for the impedimetric detection of SARS-CoV-2. The gold working electrode was modified with a double-ended thiolated poly-adenine probe, which was conjugated with magnetic Fe₃O₄@Au nanoparticles (FeO@Au-(S-polyA-S)-Au). After the loop-mediated isothermal amplification (LAMP) of viral RNA, the single-guide RNA (sgRNA), specifically bound to the SARS-CoV-2 target sequence, activates Cas12a. Cas12a then cleaved the immobilized probe. As a result, the magnetic FeO@Au nanoparticles were released and adsorbed onto the gold electrode surface, using an external magnet. This process increased the physical surface area of the gold electrode, facilitating redox ion ([Fe(CN)]) electron transfer. The decrease in the charge transfer resistance was utilized for SARS-CoV-2 detection. Our LAMP-CRISPR/Cas12a-based impedimetric biosensor, powered by FeO@Au-(S-polyA-S)-Au, demonstrated impressive capabilities, including a remarkable detection limit of 0.8 aM (0.48 copies/µL) and a linear range of 0.01 to 36.06 fM.
一种低成本、实验室制造的聚四氟乙烯微腔，配备三个电极，被开发用于 SARS-CoV-2 的阻抗检测。金工作电极用双端硫代聚腺苷酸探针进行修饰，该探针与磁性 Fe₃O₄@Au 纳米粒子（FeO@Au-(S-polyA-S)-Au）偶联。在病毒 RNA 的环介导等温扩增（LAMP）之后，与 SARS-CoV-2 靶序列特异性结合的单指导 RNA（sgRNA）激活 Cas12a。Cas12a 随后切割固定化探针。结果，磁性 FeO@Au 纳米粒子被释放并通过外部磁铁吸附到金电极表面。这一过程增加了金电极的物理表面积，促进了氧化还原离子([Fe(CN)₆])的电子转移。通过减少电荷转移电阻来检测 SARS-CoV-2。我们的基于 LAMP-CRISPR/Cas12a 的阻抗生物传感器，由 FeO@Au-(S-polyA-S)-Au 提供动力，具有令人印象深刻的性能，包括 0.8 aM（0.48 拷贝/μL）的显著检测限和 0.01 至 36.06 fM 的线性范围。
