细胞因子风暴（CS）是一种危险的免疫过度反应，伴随着干扰素-γ（IFN-γ）、白细胞介素和肿瘤坏死因子等前炎症细胞因子的显著升高。对细胞因子的敏感检测有助于研究CS的进展并诊断传染性疾病。在本研究中，我们开发了一种串联系统，将适配体、链置换扩增（SDA）、CRISPR/Cas12a和钴氢氧化物纳米片（称为Apt-SCN串联系统）作为一种信号放大平台用于IFN-γ的检测。由于亲和力较强，靶向IFN-γ与适配体结合，形成适配体-互补DNA（Apt-cDNA）双链。从Apt-cDNA双链释放的cDNA启动了SDA，生成能激活CRISPR/Cas12a的双链DNA产物。激活的CRISPR/Cas12a进一步切割带有FAM标记的单链DNA探针，阻止其附着在钴氢氧化物纳米片上并恢复荧光信号。对IFN-γ的敏感荧光分析成功地进行了，检测限度低至0.37 nM。与传统的蛋白质分析方法不同，Apt-SCN串联系统结合了多种信号放大技术，也可以适用于其他细胞因子的分析。本研究是首次利用SDA和CRISPR/Cas12a检测IFN-γ的研究，显示出在细胞因子的临床分析和CS预防方面具有巨大潜力。版权所有©2023 Elsevier B.V. 所有权利保留。

Cytokine storm (CS) is a risky immune overreaction accompanied by significant elevations of pro-inflammatory cytokines including interferon-γ (IFN-γ), interleukin and tumor necrosis factor. Sensitive detection of cytokine is conducive to studying CS progress and diagnosing infectious diseases. In this study, we developed a tandem system combining aptamer, strand displacement amplification (SDA), CRISPR/Cas12a, and cobalt oxyhydroxide nanosheets (termed Apt-SCN tandem system) as a signal-amplified platform for IFN-γ detection. Owing to the stronger affinity, target IFN-γ bound specifically to the aptamer from aptamer-complementary DNA (Apt-cDNA) duplex. The cDNA released from the Apt-cDNA duplex initiated SDA, resulting in the generation of double-stranded DNA products that could activate the trans-cleavage activity of CRISPR/Cas12a. The activated CRISPR/Cas12a further cleaved FAM-labeled single-stranded DNA probe, preventing it from adhering to the cobalt oxyhydroxide nanosheets and recovering the fluorescence signal. Sensitive fluorometric analysis of IFN-γ was successfully performed with detection limit as low as 0.37 nM. Unlike traditional protein analysis methods, Apt-SCN tandem system incorporates multiple signal amplification techniques and may also be applicable for other cytokines assay. This study was the initial study to utilize SDA and CRISPR/Cas12a to detect IFN-γ, showing great potential for cytokines clinical assay and CS prevention.Copyright © 2023 Elsevier B.V. All rights reserved.