Characterization of the Affinity Between Low Molecular Weight Targets and Their Aptamers

doi:10.13523/j.cb.20191111

China Biotechnology

2019, Vol. 39

Issue (11): 96-104 DOI: 10.13523/j.cb.20191111

Characterization of the Affinity Between Low Molecular Weight Targets and Their Aptamers

SU Yi,JIANG Ling-li,LIN Jun-sheng(

)

School of Medicine, HuaQiao University, Quanzhou 362021, China

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Abstract

Aptamers are single-stranded DNA or RNA who bind to their targets with high specificity and affinity obtained by selection from a library through SELEX. At present, there are only a few aptamers with high affinity and high specificity to low molecular weight targets all over the world. The aptamers against low molecular weight targets are difficult to be screened. Moreover, it is difficult to determine the affinity of low molecular weight targets binding with their candidate aptamers. Affinity characterization is the key step to determine whether the aptamer screening is successful or not. The existing affinity characterization methods for low molecular weight targets and their corresponding aptamers are summarized, including nanogold colorimetry, isothermal drop calorimetry, surface plasmon resonance, circular dichroism, quartz crystal microbalance, microscale thermophoresis and SYBR Green I dye detection. The advantages and disadvantages of these methods and suggestions for improvement are also discussed in order to improve the efficiency of aptamer characterization.

Key words： Aptamer Low molecular weight target Affinity characterization

Received: 04 April 2019 Published: 17 December 2019

ZTFLH:

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Corresponding Authors: Jun-sheng LIN E-mail: junshenglin@hqu.edu.cn

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	Yi SU
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	Jun-sheng LIN

Cite this article:

SU Yi,JIANG Ling-li,LIN Jun-sheng. Characterization of the Affinity Between Low Molecular Weight Targets and Their Aptamers. China Biotechnology, 2019, 39(11): 96-104.

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https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20191111 OR https://manu60.magtech.com.cn/biotech/Y2019/V39/I11/96

方法	基本原理	靶标案例	K_d值	适用范围	优势	不足	参考文献
纳米金法	普通比色法:纳米金聚集与分散状态颜色变化	啶虫脒	5nmol/L	靶标分子本身为无色	现象明显易制备,操作简单,耗时短	普适性不强,部分小分子靶标甚至会促进适配体更加稳定地吸附在纳米金上而出现相反的检测结果	[4]
		妥布霉素	23.3nmol/L				[5]
		钾离子	0.42nmol/L				[6]
		三聚氰胺	14.9nmol/L				[7]
	荧光猝灭活性	赭曲霉毒素A	5nmol/L				[8]
	荧光猝灭活性	雌二醇	0.48nmol/L				[9]
	过氧化氢酶样活性	玉米烯酮	10ng/ml	[10]
	过氧化氢酶样活性	磺胺二甲氧嘧啶	10ng/ml	[11]
ITC	检测靶标与适配体结合时热量变化	L-酪氨酰胺	16.8μmol/L	范围较广	精确度高,实时连续监测,可直接测得K_d值	用样量较大,低通量,需专人操作	[12]
		替硝唑	1nmol/L				[13]
		孔雀石绿	(50±20)nmol/L				[14]
SPR	检测适配体与靶标结合前后光折射率变化	妥布霉素	(1.24±0.03) nmol/L	需固定适配体或者靶标	灵敏度高,实时性号,芯片可再生	小分子检测信号低,小分子难固定、低通量	[15]
		黄曲霉毒素1	0.94ng/ml				[16]
		氟乙酰胺	1μmol/L				[17]
CD	检测适配体与靶标结合前后左旋及右旋光变化	25-羟基维生素D3	11nmol/L	适配体与靶标结合前后结构变化明显	快速、高效反映适配体结构变化	样品制备时纯度等要求高、前处理复杂	[18]
QCM	适配体与靶标结合前后重量变化,转化为频率变化检测	茶碱	0.526μmol/L	需固定适配体或者靶标	灵敏度高、实时性强	检测易受温度、湿度和震动等环境因素影响	[19]
QCM	适配体与靶标结合前后重量变化,转化为频率变化检测	L -酪氨酰胺	(181±27) μmol/L	需固定适配体或者靶标	灵敏度高、实时性强	检测易受温度、湿度和震动等环境因素影响	[20]
MST	靶标与适配体在毛细管中通过温度梯度热运动,检测反应体系中荧光分布的变化	17β-雌二醇	98nmol/L	待测物分子大小不限	样品量较少、可测量天然状态下适配体与靶标之间的结合	适配体需要修饰荧光,荧光寿命短	[21]
MST	靶标与适配体在毛细管中通过温度梯度热运动,检测反应体系中荧光分布的变化	ATP	(31±3) μmol/L	待测物分子大小不限	样品量较少、可测量天然状态下适配体与靶标之间的结合	适配体需要修饰荧光,荧光寿命短	[22]
SGI	SYBR Green I绿色荧光染料,检测结合前后荧光变化	ATP	23.4nmol/L	靶标无荧光	简单、易操作	检测范围小、低通量	[23]
		赭曲霉毒素A	(370±250) nmol/L				[24]
		四环素	0.10μg/ml				[25]
		呋喃酮	(1.1±0.4) μmol/L				[26]

Table 1 Summary of affinity detection methods for small molecular targets and their aptamers


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