CN114509421B - 一种密接有序的表面增强拉曼基底及其制备方法 - Google Patents
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Abstract
本发明公开了一种密接有序的表面增强拉曼基底及其制备方法,金纳米粒子通过桥链与DNA墨水链的连接,高密度接触且均匀有序排列在金膜衬底上。采用不同长度的DNA墨水链可实现不同高度的纳米粒子有序分层。通过预定DNA墨水链的位置可实现同层纳米粒子的紧密堆积,从而获得高密度有序排列的单层或多层的纳米粒子拉曼基底。本发明提出的拉曼基底能产生比较密集且均匀的热点,可提供高灵敏度和高精度的拉曼光谱检测。
Description
技术领域
本发明涉及纳米技术领域,涉及一种密接有序的表面增强拉曼基底及其制备方法。
背景技术
表面增强拉曼散射(Surface-Enhanced Raman Scattering,SERS)是一种灵敏的化学物质光谱分析技术,其主要是利用贵金属纳米颗粒的局域表面等离子体共振来极大地增强待测物分子的拉曼散射信号。目前其已广泛应用于生物、化学和医学等领域。
SERS基底的制备是SERS研究中的重点和热点。为了达到更高的检测限和灵敏度,具有更多热点的SERS基底研究受到越来越多的关注。传统的纳米阵列SERS基底都是通过在高浓度贵金属纳米粒子溶胶中直接进行自组装,其纳米粒子间隔大且排布不均匀,往往会造成“热点”不均匀,从而使得拉曼探测技术灵敏度和复现性大大降低。因此,获得有序紧密接触且热点均匀的SERS基底仍然是一个挑战。
发明内容
针对现有技术中所存在的不足,本发明提出DNA墨水法实现密接有序的金纳米阵列表面增强拉曼散射基底,该基底具有高密度且分布均匀的“热点”,该SERS基底解决了低浓度下拉曼检测的高灵敏度和高复现性问题。
本发明的上述技术目的是通过以下技术方案得以实现的:
所有的纳米粒子由DNA墨水链来连接和控制。其中不同层的纳米粒子采用DNA墨水链的长度来控制,同层的纳米粒子采用DNA墨水链的位置来控制,从而获得高密度有序排列的单层或多层的纳米粒子拉曼基底。
具体实现步骤如下:
步骤一、设计DNA墨水图案。设计含有预定墨水链位置的图形;
步骤二、制备DNA墨水图案。通过将预设的长链、短链和固定链按一定比例混合,形成有序排列的墨水图形。不同的修饰短链有序排布在设计位置形成墨水链。
步骤三、DNA墨水图案固定在金膜上。墨水图案通过上一步骤形成的墨水链下方硫醇键固定在金膜表面,墨水链上方形成不同长度的纳米粒子连接点;
步骤四、洗涤掉没有墨水链的图案结构,暴露出DNA墨水链;
步骤五、金纳米粒子表面硫醇化,使其与寡核苷酸连接。
步骤六、金纳米粒子与金膜形成表面增强拉曼基底(SERS)。金纳米粒子通过桥链实现与DNA墨水链的连接。不同层DNA墨水链的长度不同,金纳米粒子有序排布在不同高度,形成纵向和横向都紧密接触有序排布的SERS基底。
与现有技术相比,本发明具有以下优良性质:
一、由于采用了不同长度的DNA墨水链,可以实现纳米粒子纵向分层有序排列。
二、通过预设DNA墨水链的横向位置,可以实现纳米粒子横向高密度有序排列。
三、由于采用纵向和横向高密度有序排列,可以实现更多的拉曼“热点”。
附图说明
图1为设计的DNA墨水图案;
图2为DNA墨水链与金膜表面连接示意图;
图3为DNA化的金纳米粒子与桥链连接示意图;
图4为三维有序排列的SERS基底截面示意图。
具体实施方式
下面结合附图及实施例对本发明中的技术方案进一步说明。
在本实施例中,需要制备的是具有两层纳米粒子的拉曼基底,每层6×9个纳米粒子,金纳米粒子大小为30nm,纵向纳米粒子的间距为2nm,同层横向纳米粒子的间距为2nm。
为实现上述拉曼基底,采取以下步骤:
第一步、设计DNA墨水图案。通过计算机设计出500nm×500nm DNA墨水框架图,框架图案由一条长链和若干短链混合折叠而成,如图1所示。四周设计四个固定链固定整个DNA框架与金膜相连。所有的短链对应的DNA序列将由计算机设计出,数目约4700。中间部分设计成墨水链位置,即纳米粒子所在位置。墨水链设计为两种长度,短的将形成第一层纳米粒子(白圆圈加内×表示),长的将形成第二层纳米粒子(黑圆圈加内×表示)。墨水链的数目为两层之和,共108个。长墨水链比短墨水链高27nm,长墨水链与短墨水链的间距为32nm。此外,组成DNA墨水链的5′端都需要设计成面对着结构的同一平面。
第二步、制备DNA墨水图案。长链采用长线M13p8噬菌体,普通短链根据计算机设计的特定序列进行合成,其中112个进行改性,108个是用于墨水链,4个用于固定链。然后将长链、短链(包括固定链)按1∶10的摩尔比例进行混合,形成DNA墨水图案。上述过程中,DNA墨水链的改性是用经硫醇改性的短链取代框架结构中预定位置的短链而形成,具体位置为在链的5′端引入低聚胸苷基团(9或10碱基)。固定链和墨水链改性相同,区别在于墨水链间距排布紧密,主要用来连接纳米粒子而固定链分布在墨水框架的四角。
第三步、将DNA墨水框架结构固定在金膜表面。由于墨水链5′端可与金膜表面反应形成硫醇金键具有较强的连接作用,故DNA墨水框架结构可固定在金膜表面。
第四步、洗涤掉没有墨水链的图案结构,暴露出DNA墨水链。由于墨水链5′端可与金膜表面反应形成硫醇金键具有较强的连接作用,而DNA墨水框架结构与金膜表面吸附力微弱,因此采用氢氧化钠处理框架结构使其变性,洗涤掉不含墨水链的图案,暴露出长度不同的DNA墨水链,如图2所示。
第五步、金纳米粒子的改性。将硫醇-寡核苷酸链和胶体金溶液室温下混合并搅拌均匀,经过硫化过程实现金纳米粒子和寡核苷酸链的连接。所用硫醇-寡核苷酸链序列为A-CG:5’-TGACTCAATGACTCGTTTTTTTTTT-3’-磷酸盐-(CH2)3-SH)。为了实现寡核苷酸链和墨水链上端的连接,在硫醇-寡核苷酸链序列末端引入桥链(一种短链),如图3所示。
第六步、实现金纳米粒子与金膜的连接。桥链两端碱基分别与墨水链上端和寡核苷酸链互补,基于碱基互补配对原则,实现了金膜和纳米粒子间的连接。短墨水链与金纳米粒子形成第一层,长墨水链与金纳米粒子形成第二层,形成密接有序的具有三维热点的表面增强拉曼基底,如图4所示。
最后说明的是,以上实施例仅用以说明本发明的技术方案而非限制,尽管参照较佳实施例对本发明进行了详细说明,本领域的普通技术人员应当理解,可以对本发明的技术方案进行修改或者等同替换,而不脱离本发明技术方案的宗旨和范围,其均应涵盖在本发明的权利要求范围当中。
Claims (10)
1.一种密接有序的表面增强拉曼基底,其特征在于:
1)其结构由衬底、金膜、DNA墨水图案、单层或多层金纳米粒子组成;
2)制备方法包括以下步骤:
步骤一、设计DNA墨水图案,设计含有预定墨水链位置的图形;步骤二、制备DNA墨水图案,通过将预设的长链、短链和固定链按一定比例混合,形成有序排列的墨水图形,不同的修饰短链有序排布在设计位置形成墨水链;步骤三、DNA墨水图案固定在金膜上,墨水图案通过上一步骤形成的墨水链下方硫醇键固定在金膜表面,墨水链上方形成不同长度的纳米粒子连接点;步骤四、洗涤掉没有墨水链的图案结构,暴露出DNA墨水链;步骤五、金纳米粒子表面硫醇化,使其与寡核苷酸连接;步骤六、金纳米粒子与金膜形成表面增强拉曼基底(SERS),金纳米粒子通过桥链实现与DNA墨水链的连接,不同层DNA墨水链的长度不同,金纳米粒子有序排布在不同高度,形成纵向和横向都紧密接触有序排布的SERS基底。
3)通过在DNA墨水图案中修改墨水链纵向及横向排列,形成任意结构的拉曼热点。
2.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:所述金纳米粒子为多层时,纳米粒子高密度接触且有序排列,不同层金属纳米粒子与DNA墨水链由不同长度DNA链连接。
3.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:所述金纳米粒子范围为3nm-30nm。
4.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:所述金膜衬底材质包括玻璃、硅、氮化硅、二氧化硅。
5.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:所述衬底上金膜的厚度为10nm-50nm。
6.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:所述DNA墨水图案由短链、长链和固定链组成,且按一定比例混合,所述短链包括包括普通短链和修饰短链。
7.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:所述金纳米粒子为多层时,不同层纳米粒子间距可通过不同DNA墨水链的长度来控制,最小间距为1nm。
8.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:纳米粒子间距通过修改DNA墨水链连接点控制,最小间距为1nm。
9.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:基底上的纳米粒子排布成任意的二维或三维图案。
10.如权利要求1所述的密接有序的表面增强拉曼基底,其特征在于:所述金纳米粒子为多层时,金纳米粒子有序排布在不同高度,形成纵向和横向都紧密接触有序排布的SERS基底。
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