基于核酸探针的荧光各向异性检测方法  

Fluorescence anisotropy detection methods based on nucleic acid probes

作　　者：谢佳丽 秦雨 吴竹莲[1] 甄淑君[1] Jiali Xie;Yu Qin;Zhulian Wu;Shujun Zhen(College of Chemistry and Chemical Engineering,Southwest University,Chongqing 400715,China)

机构地区：[1]西南大学化学化工学院,重庆400715

出　　处：《科学通报》2024年第26期3900-3913,共14页Chinese Science Bulletin

基　　金：国家自然科学基金(22322409,21974109)资助。

摘　　要：荧光各向异性法是一种均相检测法,具有准确度高、可实现高通量检测的优点.近年来,基于核酸探针的荧光各向异性法已经被广泛应用于生物分子的检测.当检测溶液温度和黏度一定时,荧光各向异性值(r)与荧光分子旋转速度成反比.然而传统的荧光各向异性法由于大多数靶标分子的质量(或体积)小,旋转速度快,不易产生明显的荧光各向异性差异,导致灵敏度较低.本文详细阐述了不同信号放大方式来提高基于核酸探针的荧光各向异性法灵敏度的策略.我们相信,在高灵敏度的方法建立之后,荧光各向异性法会进入临床应用阶段.Nucleic acid probe(NAP)-based fluorescence anisotropy(FA)is an excellent biosensing method due to its high throughput,homogeneous detection ability,and accuracy.FA is a ratio determination method in which the FA value(r)is inversely proportional to the rotational speed of the fluorophore modified NAP when the temperature and viscosity of the detection solution are constant.Based on the above,the FA method has been widely utilized for detecting various biomolecules.However,the sensitivity of traditional FA approaches is limited,mostly due to the following two factors:(1)The small volume(or mass)and fast rotation speed of most target molecules make it difficult to produce significant changes in FA,resulting in poor detection sensitivity;and(2)the large volume(or mass)of the NAPs causes the background signal to be high,resulting in the change in r after addition of the target to be insignificant,thereby leading to poor sensitivity.In this review,we describe in detail the strategies for different signal amplification methods to improve the sensitivity of NAP–based FA methods.To enhance the sensitivity of FA,one strategy is to use enhancement materials of an appropriate volume(or mass)to improve the FA signal.Another is to reduce the background signal by cutting NAPs into fragments using different enzymes.As a biomacromolecule,protein not only has a large volume(or mass),but it also exhibits good biocompatibility,making it a common FA amplification material.Unfortunately,the suitability of this protein is limited,as it becomes unstable when heated and complex to operate during use.At the same time,inorganic nanomaterials,such as metal nanoparticles,carbon nanomaterials,and transition metal nanomaterials,offer advantages such as good thermal stability and easy synthesis and surface modification;as a result,they are often utilized to enhance FA signals.However,most inorganic nanomaterials have uncontrollable sizes and strong fluorescence quenching capabilities,which can reduce the accuracy of FA.DNA nanostructures are

关 键 词：核酸探针 荧光各向异性法 信背比 信号放大 

分 类 号：Q503[生物学—生物化学] O657.3[理学—分析化学]