CN116448847B - 纸基光电化学生物传感器的制备及在戊唑醇检测中的应用 - Google Patents
纸基光电化学生物传感器的制备及在戊唑醇检测中的应用 Download PDFInfo
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Abstract
本发明公开了一种纸基光电化学生物传感器的制备及在戊唑醇检测中的应用。通过于色谱纸上印制疏水蜡图案并在亲水区生长Fe2O3及Ag纳米粒子,并修饰底物探针和识别链,借助目标物特异性识别CuInS2修饰的识别链,导致电极光电流强度发生改变,实现水体中戊唑醇的快速、灵敏检测。
Description
技术领域
本发明涉及一种纸基光电化学生物传感器的制备方法,更具体地说是一种可以用于水体中戊唑醇快速检测传感平台的构建。
背景技术
戊唑醇是一种能有效控制水系统或果蔬表面细菌、真菌及藻类的化学制农药,其可以保护植物避免真菌损害,保证农业稳产、高产,满足国民对农副产品的巨大需求。然而,农业中过渡和违规使用的戊唑醇,可通过地表径流及沉降途径进入水环境,进而对地表水及地下水等造成污染,对人类健康产生威胁。因此,建立简单、快速高选择性戊唑醇检测方法成为广大科研工作者重大攻关课题之一。
现有戊唑醇检测技术,通常包括荧光法、质谱法、高效液相色谱法、液-质联用技术等。以上检测技术准确度高,选择性好,然而存在检测仪器不便携及检测成本高等问题,因此难于完成水体环境中戊唑醇现场快速检测。
微流控纸芯片,兼具样品采集、富集、分离及检测等功能,且芯片制备成本低,可批量加工,质量轻、便携等,由此适合于大众化现场及时检测(POCT)。以纸芯片作为戊唑醇识别富集平台,结合光电化学技术高的检测精度,有望解决现有检测技术中存在的仪器不便携、检测成本高等问题,更好地推动生态环境保护高质量发展,提高人民生活质量。
发明内容
本发明要解决的技术问题是制备一种纸基光电化学生物传感器,以实现水体环境中戊唑醇的灵敏检测。
为了解决上述技术问题,本发明是通过以下措施来实现的:一种纸基光电化学生物传感器的制备方法,其特征是包括以下步骤:
(1)选用色谱纸作为基底,利用裁纸刀将色谱纸裁剪成A4纸大小,随后利用蜡打印机于A4纸上印制疏水蜡图案;
(2)将印有疏水蜡图案的A4纸放于烘箱中,120 °C下加热使蜡溶化,形成疏水墙,样式如附图1所示,其中白色区域为亲水区,灰色部分为疏水区;
(3)对(2)中亲水区进行功能化,制备Fe2O3及Ag纳米粒子功能化的纸芯片电极:
所述Fe2O3及Ag纳米粒子功能化的纸芯片电极的制备,包括以下步骤:在纸芯片上重复三次滴加100 μL的Ag纳米粒子,室温下干燥,随后将纸芯片放于盛有浓度为100 mM的AgNO3及10 mM抗坏血酸混合液中,室温下以2000 rpm转速下磁力搅拌反应5分钟,而后将纸芯片取出,二次水冲洗表面,即可获得Ag纳米粒子功能化的纸芯片电极;随后,以Ag纳米粒子功能化的纸芯片电极作为生长基底,于其表面进一步生长Fe2O3纳米层:将Ag纳米粒子功能化的纸芯片放于10 mL浓氨水、20 mL水、5 mL乙醇及6 mL浓度为18 mM的单宁酸混合液中,而后,向混合液中加入0.5 mL的甲醛溶液,35 °C下反应16 h,随后,将纸芯片取出,二次水冲洗表面,室温下干燥;将纸芯片放于50 mL高压釜中,向高压釜内加入体积为30 mL、浓度为0.07 M的Fe(NO3)3溶液,110 °C下反应2 h,将纸芯片取出,二次水冲洗表面后,60 °C下干燥,即可获得Fe2O3及Ag纳米粒子功能化的纸芯片电极,定义为PWE-Ag-Fe2O3;
(4)将一定浓度的氨基标记的底物探针,定义为链1,其碱基序列如核苷酸序列表所示,加入到步骤(3)所得PWE-Ag-Fe2O3表面,室温下反应45分钟,随后用磷酸盐缓冲溶液清洗电极表面;
所述氨基标记的底物探针,其特征是其5'端标记有氨基;
(5)将一定浓度的CuInS2修饰的且带有氨基标记的识别链,定义为链2,其碱基序列如核苷酸序列表所示,加入到步骤(4)所得电极表面,室温下反应2 h,随后用磷酸盐缓冲溶液冲洗电极表面,完成链1和链2间杂交链反应;
所述CuInS2修饰的识别链的制备,包括以下步骤:将60 mg的CuInS2纳米材料溶于50 mL水中,加入浓度为0.15 mM、体积为1 mL的链2,35 °C下反应12 h,将所得产物8000rpm转速下离心10分钟,最后将产物分散在2 mL无霉无菌水溶液中,即可得到CuInS2修饰的链2;
所述氨基标记的识别链,其特征是其5'端标记有氨基;
(6)移取20 μL不同地区采集到的水样滴加到步骤(5)所得电极表面,室温下反应40分钟,而后用二次水冲洗电极3次,将电极室温下干燥;
(7)将纸芯片作为工作电极,以白光作为激发光源,采用三电极系统记录光电流强度,绘制光电流强度与戊唑醇浓度关系曲线,完成水体环境中戊唑醇的灵敏检测。
本发明的有益效果:
(1)该制备方法简单、通用。
(2)所制备的PWE-Ag-Fe2O3纳米材料展现出良好的光电转换效率,且制备成本低廉。
(3)该方法可完成复杂水样中戊唑醇的选择性识别和灵敏检测。
附图说明
图1:印有蜡图案的A4纸;
具体实施方式
为了更好地理解本发明,下面通过实施例进一步阐明本发明的内容,但本发明不仅仅局限于下面的实施例。
实施例1
一种纸基光电化学生物传感器的制备,具体制备工艺如下:
(1)选用色谱纸作为基底,利用裁纸刀将色谱纸裁剪成A4纸大小,随后利用蜡打印机于A4纸上印制疏水蜡图案;
(2)将印有疏水蜡图案的A4纸放于烘箱中,120 °C下加热使蜡溶化,形成疏水墙,其中白色区域为亲水区,灰色部分为疏水区;
(3)对(2)中亲水区进行功能化,制备Fe2O3及Ag纳米粒子功能化的纸芯片电极:
所述Fe2O3及Ag纳米粒子功能化的纸芯片电极的制备,包括以下步骤:在纸芯片上重复三次滴加100 μL的Ag纳米粒子,室温下干燥,随后将纸芯片放于盛有浓度为100 mM的AgNO3及10 mM抗坏血酸混合液中,室温下以2000 rpm转速下磁力搅拌反应5分钟,而后将纸芯片取出,二次水冲洗表面,即可获得Ag纳米粒子功能化的纸芯片电极;随后,以Ag纳米粒子功能化的纸芯片电极作为生长基底,于其表面进一步生长Fe2O3纳米层:将Ag纳米粒子功能化的纸芯片放于10 mL浓氨水、20 mL水、5 mL乙醇及6 mL浓度为18 mM的单宁酸混合液中,而后,向混合液中加入0.5 mL的甲醛溶液,35 °C下反应16 h,随后,将纸芯片取出,二次水冲洗表面,室温下干燥;将纸芯片放于50 mL高压釜中,向高压釜内加入体积为30 mL、浓度为0.07 M的Fe(NO3)3溶液,110 °C下反应2 h,将纸芯片取出,二次水冲洗表面后,60 °C下干燥,即可获得Fe2O3及Ag纳米粒子功能化的纸芯片电极,定义为PWE-Ag-Fe2O3;
(4)将一定浓度的氨基标记的底物探针,定义为链1,加入到步骤(3)所得PWE-Ag-Fe2O3表面,室温下反应45分钟,随后用磷酸盐缓冲溶液清洗电极表面;
所述氨基标记的底物探针,其特征是其5'端标记有氨基;
(5)将一定浓度的CuInS2修饰的且带有氨基标记的识别链,定义为链2,加入到步骤(4)所得电极表面,室温下反应2 h,随后用磷酸盐缓冲溶液冲洗电极表面,完成链1和链2间杂交链反应;
所述CuInS2修饰的识别链的制备,包括以下步骤:将60 mg的CuInS2纳米材料溶于50 mL水中,加入浓度为0.15 mM、体积为1 mL的链2,35 °C下反应12 h,将所得产物8000rpm转速下离心10分钟,最后将产物分散在2 mL无霉无菌水溶液中,即可得到CuInS2修饰的链2;
所述氨基标记的识别链,其特征是其5'端标记有氨基;
(6)移取20 μL不同地区采集到的水样滴加到步骤(5)所得电极表面,室温下反应40分钟,而后用二次水冲洗电极3次,将电极室温下干燥;
(7)将纸芯片作为工作电极,以白光作为激发光源,采用三电极系统记录光电流强度,绘制光电流强度与戊唑醇浓度关系曲线,完成水体环境中戊唑醇的灵敏检测。
Claims (1)
1.一种纸基光电化学生物传感器的制备方法,其特征在于该方法的制备步骤如下:
(1)选用色谱纸作为基底,利用裁纸刀将色谱纸裁剪成A4纸大小,随后利用蜡打印机于A4纸上印制疏水蜡图案;
(2)将印有疏水蜡图案的A4纸放于烘箱中,120 °C下加热使蜡溶化,形成疏水墙,其中白色区域为亲水区,灰色部分为疏水区;
(3)对(2)中亲水区进行功能化,制备Fe2O3及Ag纳米粒子功能化的纸芯片电极:
所述Fe2O3及Ag纳米粒子功能化的纸芯片电极的制备,包括以下步骤:在纸芯片上重复三次滴加100 μL的Ag纳米粒子,室温下干燥,随后将纸芯片放于盛有浓度为100 mM的AgNO3及10 mM抗坏血酸混合液中,室温下以2000 rpm转速下磁力搅拌反应5分钟,而后将纸芯片取出,二次水冲洗表面,即可获得Ag纳米粒子功能化的纸芯片电极;随后,以Ag纳米粒子功能化的纸芯片电极作为生长基底,于其表面进一步生长Fe2O3纳米层:将Ag纳米粒子功能化的纸芯片放于10 mL浓氨水、20 mL水、5 mL乙醇及6 mL浓度为18 mM的单宁酸混合液中,而后,向混合液中加入0.5 mL的甲醛溶液,35 °C下反应16 h,随后,将纸芯片取出,二次水冲洗表面,室温下干燥;将纸芯片放于50 mL高压釜中,向高压釜内加入体积为30 mL、浓度为0.07 M的Fe(NO3)3溶液,110 °C下反应2 h,将纸芯片取出,二次水冲洗表面后,60 °C下干燥,即可获得Fe2O3及Ag纳米粒子功能化的纸芯片电极,定义为PWE-Ag-Fe2O3;
(4)将一定浓度的氨基标记的底物探针,定义为链1,加入到步骤(3)所得PWE-Ag-Fe2O3表面,室温下反应45分钟,随后用磷酸盐缓冲溶液清洗电极表面;
所述氨基标记的底物探针,其特征是其5'端标记有氨基;
(5)将一定浓度的CuInS2修饰的且带有氨基标记的识别链,定义为链2,加入到步骤(4)所得电极表面,室温下反应2 h,随后用磷酸盐缓冲溶液冲洗电极表面,完成链1和链2间杂交链反应;
所述CuInS2修饰的识别链的制备,包括以下步骤:将60 mg的CuInS2纳米材料溶于50 mL水中,加入浓度为0.15 mM、体积为1 mL的链2,35 °C下反应12 h,将所得产物8000 rpm转速下离心10分钟,最后将产物分散在2 mL无霉无菌水溶液中,即可得到CuInS2修饰的链2;
所述氨基标记的识别链,其特征是其5'端标记有氨基;
(6)移取20 μL不同地区采集到的水样滴加到步骤(5)所得电极表面,室温下反应40分钟,而后用二次水冲洗电极3次,将电极室温下干燥;
(7)将纸芯片作为工作电极,以白光作为激发光源,采用三电极系统记录光电流强度,绘制光电流强度与戊唑醇浓度关系曲线,完成水体环境中戊唑醇的灵敏检测。
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