清脆的
生物传感器
核酸酶
检出限
辣根过氧化物酶
适体
分子生物学
化学
DNA
纳米技术
材料科学
生物
色谱法
酶
生物化学
基因
作者
Yi Li,Fei Deng,Ewa M. Goldys
标识
DOI:10.1016/j.snb.2021.130533
摘要
The collateral cleavage function is only a corollary of programmable nuclease activity of certain Cas effectors, such as Cas12 and Cas13, but it can be utilised to amplify fluorescence signals in various CRISPR/Cas-based biosensing systems. In this work, this special signal amplification capability of CRISPR/Cas12a ribonucleoproteins has been employed to increase the sensitivity of a broad class of commercial ELISA systems with undisclosed chemistry except for the use of horseradish peroxidase (HRP), a common signal reporting molecule. We demonstrated that such ELISA systems with HRP on the detection antibody, can be amplified by 2 orders of magnitude using an example commercial IFN-γ ELISA kit where the detection limit was decreased from 312.5 pg/mL to 1.2 pg/mL. The detection range was simultaneously increased from 2 orders to 3 orders of magnitude. Our CRISPR/Cas12a-based ELISA Sensitivity Amplifier (CES-Amplifier) approach is based on a hybrid single strand DNA oligo and antibody conjugate targeting the HRP enzyme. The CES-Amplifier can be directly integrated into commercial ELISA kits to replace their original last step, without any additional changes of the ELISA kit reagents or setup. In this way, our CES-Amplifier provides a versatile and affordable approach for expanding CRISPR/Cas-based biosensing to a wide range of non-nucleic acid analytes.
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