基于核酸适体的新型液晶生物传感用于检测血小板源性生长因子BB

李霞等
摘 要 利用核酸适体与其靶分子具有高度特异性结合的原理,建立了基于液晶取向改变的以核酸适体为捕获探针用于检测血小板源性生长因子BB(Plateletderived growth factor BB, PDGFBB)分子的新型液晶生物传感方法。核酸适体通过戊二醛偶联固定在3氨丙基三乙氧基硅烷/N,N二甲基N十八烷基[3(三甲氧基硅基)丙基]氯化铵[(3Aminopropyl)trimethoxysilane/N,NdimethylNoctadecyl(3aminopropyl)trimethoxysilyl chloride, APTES/DMOAP]混合自组装的传感基底表面,当靶分子PDGFBB存在时,可与核酸适体发生特异性作用结合于传感基底表面,根据生物分子的空间尺寸效应能诱导液晶分子取向发生变化,从而引起光学信号的亮度和色彩发生改变,实现对PDGFBB的快速检测。本方法具有操作简单、选择型好、灵敏度高的特点,在PDGFBB浓度为5 nmol/L时仍可观察到明显的光学信号变化。关键词 核酸适体; 血小板源性生长因子BB; 液晶生物传感; 自组装膜
20131112收稿;20140111接受
本文系国家自然科学基金项目(Nos. 21175037, 21277042, J1210040)和湖南省科研条件创新专项重点项目(No. 2011TT1004)资助
* Email: zywu@hnu.edu.cn 1 引 言
液晶生物传感是基于液晶分子在功能膜表面排列取向发生变化,改变液晶分子对光线的折射能力,将界面上产生的反应变化信息转换成光学信号变化,实现对目标物的检测[1,2]。与传统的分析检测方法相比,液晶生物传感无需采用外源性标记物修饰,在自然光下就能进行检测,所呈现的光学信号可用肉眼观察,具有操作简单、响应快速、易于实现微型化和阵列化等特点,使其在生命科学、临床医学和环境研究上具有广泛的应用前景。蛋白质作为机体细胞的重要组成部分,是人体组织更新和修补的主要原料。它对调节生理功能,维持新陈代谢都起着重要作用。因此,蛋白质检测是目前临床诊断以及化学与生物研究上的一个重要课题[3~5]。
本研究基于大分子物质,特别是蛋白质分子的空间尺寸效应能有效干扰液晶分子有序排列的原理[6,7],以核酸适体为捕获探针建立了基于液晶取向改变的新型液晶生物传感方法用于检测血小板源性生长因子BB(Plateletderived growth factor BB, PDGFBB)。PDGFBB作为人体血小板中含有的一种促进细胞有丝分裂、趋化活性的生长因子[8~11],在正常细胞中含量较少,但在一些人类肿瘤细胞中却常常发生基因过表达,是癌症诊断中的一种标志物[12],因此发展对其灵敏的检测方法引起了人们很大的兴趣。本研究结合液晶生物传感的优点,建立了一种无需标记、操作简单、选择性好、成本低,响应快速的液晶生物传感检测PDGFBB的方法。 2 实验部分
2.1 仪器与试剂
References
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16 LUAN RuiBo, YANG XiaoHai, WANG KeMin. Life Science Research, 2006, 10(2): 113-117
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刘 歆, 徐根明, 郭江峰, 丁先锋, 高晓莲. 中国生物工程杂志, 2008, 28(1): 55-60
AptamerBased on Liquid Crystal Biosensor for
Detemination of Plateletderived Growth Factor BB
LI Xia, TAN Hui, LI YunLei, WU ZhaoYang*, SHEN GuoLi, YU RuQin
(State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering,
Hunan University, Changsha 410082, China)
Abstract A novel liquid crystal (LC) biosensor was developed for the detection of plateletderived growth factor BB (PDGFBB) based on the orientation changes of liquid crystals. One glass slide of the LC cell was first modified with the APTES/DMOAP ((3aminopropyl)trimethoxysilane/N,NdimethylNoctadecyl(3aminopropyl)trimethoxysilyl chloride) selfassembled monolayer (SAM) to trigger the homeotropic alignment of LC molecules and thereby produced an black background optical image under the crossed polarized light, and then the specific aptamer of PDGFBB was immobilized on SAM through glutaraldehyde crosslinking to construct a LC sensing substrate. In the presence of PDGFBB, the stable triple helix structure of PDGFBB aptamer was formed by the specific binding event and thus led to the target PDGFBB captured to the sensing substrate, making a visible optical change observed under the crossed polarized light via the huge steric effect of the PDGFBB on disrupting the LC alignment. But in the absence of PDGFBB, the low packing density of the PDGFBB aptamer had little effect on disrupting the ordered alignment to the LC and caused the black background optical image still observed under the crossed polarized light. There was an obvious optical change when the concentration of target PDGFBB was 5 nmol/L. The proposed LC biosensing method permits the labelfree detection of PDGFBB with good selectivity and high sensitivity, making them sufficiently simple and particularly useful for lowcost screening bioassay performed away from central laboratories.
Keywords Aptamer; Plateletderived growth factor BB; Liquid crystal biosensor; Selfassembled monolayer
(Received 12 November 2013; accepted 11 January 2014)
This work was supported by the National Natural Science Foundation of China (Nos.21175037, 21277042 and J1210040)
11 Hart C E, Bailey M, Curtis D A, Osborn S, Raines E, Ross R, Forstrom J W. Biochem., 1990, 29(1): 1660-172
12 Zhou C S, Jiang Y X, Hou S, Ma B, Fang X H, Li M L. Anal. Bioanal. Chem., 2006, 384(5): 1175-1180
13 Liss M, Petersen B, Wolf H, Prohaska E. Anal. Chem., 2002, 74(17): 4488-4495
14 Green L S, Jellinek D, Jenison R, Ostman A, Heldin C H, Janjic N. Biochem., 1996, 35(45): 14413-14424
15 Ruslinda A R, Penmatsa V, Ishii Y, Tajima S, Kawarada H. Analyst, 2012, 137(7): 1692-1697
16 LUAN RuiBo, YANG XiaoHai, WANG KeMin. Life Science Research, 2006, 10(2): 113-117
栾瑞波, 羊小海, 王柯敏. 生命科学研究, 2006, 10(2): 113-117
17 LIU Xin, XU GenMing, GUO JiangFeng, DING XianFeng, GAO XiaoLian. China Biotechnology, 2008, 28(1): 55-60
刘 歆, 徐根明, 郭江峰, 丁先锋, 高晓莲. 中国生物工程杂志, 2008, 28(1): 55-60
AptamerBased on Liquid Crystal Biosensor for
Detemination of Plateletderived Growth Factor BB
LI Xia, TAN Hui, LI YunLei, WU ZhaoYang*, SHEN GuoLi, YU RuQin
(State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering,
Hunan University, Changsha 410082, China)
Abstract A novel liquid crystal (LC) biosensor was developed for the detection of plateletderived growth factor BB (PDGFBB) based on the orientation changes of liquid crystals. One glass slide of the LC cell was first modified with the APTES/DMOAP ((3aminopropyl)trimethoxysilane/N,NdimethylNoctadecyl(3aminopropyl)trimethoxysilyl chloride) selfassembled monolayer (SAM) to trigger the homeotropic alignment of LC molecules and thereby produced an black background optical image under the crossed polarized light, and then the specific aptamer of PDGFBB was immobilized on SAM through glutaraldehyde crosslinking to construct a LC sensing substrate. In the presence of PDGFBB, the stable triple helix structure of PDGFBB aptamer was formed by the specific binding event and thus led to the target PDGFBB captured to the sensing substrate, making a visible optical change observed under the crossed polarized light via the huge steric effect of the PDGFBB on disrupting the LC alignment. But in the absence of PDGFBB, the low packing density of the PDGFBB aptamer had little effect on disrupting the ordered alignment to the LC and caused the black background optical image still observed under the crossed polarized light. There was an obvious optical change when the concentration of target PDGFBB was 5 nmol/L. The proposed LC biosensing method permits the labelfree detection of PDGFBB with good selectivity and high sensitivity, making them sufficiently simple and particularly useful for lowcost screening bioassay performed away from central laboratories.
Keywords Aptamer; Plateletderived growth factor BB; Liquid crystal biosensor; Selfassembled monolayer
(Received 12 November 2013; accepted 11 January 2014)
This work was supported by the National Natural Science Foundation of China (Nos.21175037, 21277042 and J1210040)
11 Hart C E, Bailey M, Curtis D A, Osborn S, Raines E, Ross R, Forstrom J W. Biochem., 1990, 29(1): 1660-172
12 Zhou C S, Jiang Y X, Hou S, Ma B, Fang X H, Li M L. Anal. Bioanal. Chem., 2006, 384(5): 1175-1180
13 Liss M, Petersen B, Wolf H, Prohaska E. Anal. Chem., 2002, 74(17): 4488-4495
14 Green L S, Jellinek D, Jenison R, Ostman A, Heldin C H, Janjic N. Biochem., 1996, 35(45): 14413-14424
15 Ruslinda A R, Penmatsa V, Ishii Y, Tajima S, Kawarada H. Analyst, 2012, 137(7): 1692-1697
16 LUAN RuiBo, YANG XiaoHai, WANG KeMin. Life Science Research, 2006, 10(2): 113-117
栾瑞波, 羊小海, 王柯敏. 生命科学研究, 2006, 10(2): 113-117
17 LIU Xin, XU GenMing, GUO JiangFeng, DING XianFeng, GAO XiaoLian. China Biotechnology, 2008, 28(1): 55-60
刘 歆, 徐根明, 郭江峰, 丁先锋, 高晓莲. 中国生物工程杂志, 2008, 28(1): 55-60
AptamerBased on Liquid Crystal Biosensor for
Detemination of Plateletderived Growth Factor BB
LI Xia, TAN Hui, LI YunLei, WU ZhaoYang*, SHEN GuoLi, YU RuQin
(State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering,
Hunan University, Changsha 410082, China)
Abstract A novel liquid crystal (LC) biosensor was developed for the detection of plateletderived growth factor BB (PDGFBB) based on the orientation changes of liquid crystals. One glass slide of the LC cell was first modified with the APTES/DMOAP ((3aminopropyl)trimethoxysilane/N,NdimethylNoctadecyl(3aminopropyl)trimethoxysilyl chloride) selfassembled monolayer (SAM) to trigger the homeotropic alignment of LC molecules and thereby produced an black background optical image under the crossed polarized light, and then the specific aptamer of PDGFBB was immobilized on SAM through glutaraldehyde crosslinking to construct a LC sensing substrate. In the presence of PDGFBB, the stable triple helix structure of PDGFBB aptamer was formed by the specific binding event and thus led to the target PDGFBB captured to the sensing substrate, making a visible optical change observed under the crossed polarized light via the huge steric effect of the PDGFBB on disrupting the LC alignment. But in the absence of PDGFBB, the low packing density of the PDGFBB aptamer had little effect on disrupting the ordered alignment to the LC and caused the black background optical image still observed under the crossed polarized light. There was an obvious optical change when the concentration of target PDGFBB was 5 nmol/L. The proposed LC biosensing method permits the labelfree detection of PDGFBB with good selectivity and high sensitivity, making them sufficiently simple and particularly useful for lowcost screening bioassay performed away from central laboratories.
Keywords Aptamer; Plateletderived growth factor BB; Liquid crystal biosensor; Selfassembled monolayer
(Received 12 November 2013; accepted 11 January 2014)
This work was supported by the National Natural Science Foundation of China (Nos.21175037, 21277042 and J1210040)
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