物联网-智慧传输-基于聚多巴胺纳米材料和酶促循环放大策略的荧光生物传感新方法研究.pdf

摘要

I

摘要

核酸、生物酶、蛋白质以及生物小分子等参与许多重要的生物学过程，生物

分子的检测平台受到了越来越多的关注。光学生物传感器由于具有灵敏度高、检

测限低、特异性好和准确度高等优异性能被广泛用于生物分子的检测。聚多巴胺

纳米材料因其合成方法简单，并且具有独特的光学性能、吸附特性、粘附性能、

力较小。当体系中引入 T4 PNK 后，T4 PNK 使 dsDNA 的 5′-末端磷酸化，产生 5′-

磷酰基末端产物，λ exo 会水解 dsDNA 的 5′-磷酰基末端产物，并释放 ssDNA。

ssDNA 上标记的荧光基团和聚多巴胺纳米管之间产生能量共振转移（FRET）效

应，荧光强度降低。基于上述策略，本体系成功建立了一种高灵敏检测 T4 PNK

及其抑制剂的荧光传感方法，检测限低至 0.01 U/mL，线性范围为 0.01 至 50 U/mL。

同时，在复杂生物环境中成功检测了 T4 PNK，表明了该方法在生物医学和药物

筛选方面具有巨大的潜力。

2、构建基于聚多巴胺纳米管（PDANTs）和核酸外切酶 I（Exo I）的循环放

大策略检测细胞色素 C（cyt C）。 聚多巴胺纳米管能够快速吸附 cyt C 的适配体，

并猝灭其标记的荧光基团。当体系中引入目标物 cyt C 后，cyt C 能特异性结合

3、构建基于聚多巴胺纳米管（PDANTs）和 脱氧核糖核酸酶 I（DNase I）辅

助靶标循环放大平台检测 microRNA。聚多巴胺纳米管能够快速吸附单链 DNA，

并猝灭其标记的荧光基团。当体系中引入目标物 microRNA 后，microRNA 能快

速结合聚多巴胺纳米管表面上吸附的互补序列，形成 RNA-DNA 双链结构，从

纳米管表面脱落，导致荧光恢复。DNase I 只作用切割 RNA-DNA 双链结构中的

摘要

DNA 链，而对 RNA 链无活性作用。释放的 microRNA 继续结合 PDANTs 表面

吸附的互补序列，实现信号放大策略。基于上述策略，本体系成功建立了简单快

速、高灵敏检测 microRNA 的方法，检测限为 0.01 nM。此外，该实验方法可以

在血清中直接进行 microRNA 样品分析，表明了该方法在临床诊断方面具有巨大

Abstract

Nucleic acids, biological enzymes, proteins, and small biological molecules are

involved in many important biological processes. The analysis and detection of

biological molecules has received more and more attention. Optical biosensors are

widely used for the detection of biomolecules due to their excellent properties such as

high sensitivity, low detection limit, good specificity and high accuracy.

Polydopamine is the material of choice for biosensing applications due to its simple

synthesis method, unique optical properties, adsorption properties, adhesion

properties, and good biocompatibility and biodegradability. Combining the

but have less affinity for dsDNA. When T4 PNK is introduced into the system, T4

PNK phosphorylates the 5′-terminus of dsDNA to produce a 5′-phosphoryl end

product. λ exo hydrolyzes the 5′-phosphoryl end product of dsDNA and releases

ssDNA. The energy resonance transfer (FRET) effect occurs between the labeled

fluorophore and PDANTs on ssDNA, and the fluorescence intensity decreases. Based

on the above strategy, this system successfully established a highly sensitive

fluorescence sensing method for the detection of T4 PNK and its inhibitors. The

detection limit of this method is 0.01 U/mL, and the linear range is 0.01 to 50 U/mL.

At the same time, this method has achieved the detection of T4 PNK in complex

systems, indicating that this method has great potential in biomedical and drug

Abstract

single-stranded exonuclease that cleaves the aptamer of cyt C off the surface of

PDANTs, releases cyt C into the next cycle, and the fluorescence signal is stronger.

Based on the above strategy, this system successfully established a highly sensitive

fluorescence sensing method for detecting cyt C. The detection limit of this method is

0.003 μM, and the linear range is 0.01-100 μM. This method enables the detection of

cyt C in complex systems, indicating that the method can be used to monitor cell

apoptosis to study certain cell-level diseases.

3. Construction of a circular amplification platform based on polydopamine

nanotubes (PDANTs) and DNase I assisted target detection to detect microRNAs.

PDANTs can rapidly adsorb DNA probes and quench their fluorescence. When the

adsorbed on the surface of PDANTs to implement a signal amplification strategy.

Based on the above strategy, the system has successfully established a highly sensitive

method for detecting microRNAs with a detection limit as low as 0.01 nM. This

method has the advantages of simple and fast, high sensitivity and low detection limit.

In addition, the experimental method can be used to analyze microRNA samples

directly in serum, indicating that the method has great potential for clinical diagnosis.

Keywords: Fluorescence sensing method; Polydopamine; Energy resonance transfer;

Cyclic amplification; DNase I; Exonuclease I; T4 PNK; Inhibitor; Cytochrome C;

MicroRNA.

目录

V

1.2 荧光生物传感器............................................................................................. 7

1.4 立题依据与主要内容................................................................................... 16