Herein, a novel electrochemiluminescence (ECL) biosensor was proposed for ultrasensitive detection of laminin (LN), in which DNA dendrimer (D) as a promising nanocarrier for luminophore and DNA nanomachine as tactic for target recycling. The DNA dendrimer was synthesized by hybridization between sense and its antisense Y-shaped DNAs which were formed via reaction between single-stranded DNA (ssDNA) with a thiol group at the 5′-end and a synthesized trimeric cross-linker of tris(2-maleimidoethyl)amine. This dendrimer provided abundant double-stranded DNA (dsDNA) to achieve high loading efficiency for ECL luminophore. Simultaneously, N-(aminobutyl)-N-(ethylisoluminol) (ABEI) was conjugated with doxorubicin (Dox, a intercalator of dsDNA) to form the ECL indicator (Dox-ABEI) which could effectively intercalate DNA dendrimer to form the ECL probe (D-Dox-ABEI). Subsequently, a DNA nanomachine activated by target protein was involved to obtain numerous output ssDNA (S2) which was amplified by exonuclease III-assisted recycle and immobilized on ssDNA (S1)-modified sensing electrode surface via complementation. Thereby, abundant D-Dox-ABEI probes were captured by S2 to construct the biosensor for target protein detection. The proposed ECL biosensor realized the ultrasensitive detection of LN with a linear range from 0.1 pg/mL to 100 ng/mL and a low detection limit of 0.0661 pg/mL. Impressively, the application of this ECL biosensor would provide the great potential for analysis of other proteins, revealing a new avenue for early diagnosis and the prognosis evaluation of various diseases.

本文提出了一种新型的电化学发光（ECL）生物传感器用于层粘连蛋白（LN）的超灵敏检测，其中DNA树状大分子（D）作为有希望的发光体纳米载体，而DNA纳米机作为靶标回收策略。DNA树枝状大分子是通过有义和其反义Y形DNA杂交而合成的，Y形DNA是通过单链DNA（ssDNA）在5'端带有硫醇基与合成的tris（2）三聚交联剂之间的反应形成的-马来酰亚胺基乙胺）。该树枝状大分子提供了丰富的双链DNA（dsDNA），以实现ECL发光体的高加载效率。同时，将N-（氨基丁基）-N-（乙基异鲁米诺）（ABEI）与阿霉素（Dox，dsDNA插入剂形成ECL指示剂（Dox-ABEI），可有效插入DNA树枝状大分子形成ECL探针（D-Dox-ABEI）。随后，由靶蛋白激活的DNA纳米机器参与以获得大量输出ssDNA（S2），其通过核酸外切酶III辅助循环进行扩增，并通过互补固定在ssDNA（S1）修饰的传感电极表面上。由此，S2捕获了丰富的D-Dox-ABEI探针以构建用于靶蛋白检测的生物传感器。提出的ECL生物传感器实现了LN的超灵敏检测，线性范围从0.1 pg / mL到100 ng / mL，检测下限为0.0661 pg / mL。令人印象深刻的是，这种ECL生物传感器的应用将为分析其他蛋白质提供巨大的潜力，