A "signal-on" chemiluminescence biosensor was established for detecting thrombin. The thrombin aptamer1-functionalized magnetic sodium alginate (Malg-Apt1) hydrogel was synthesized by physical interaction between sodium alginate and Ca, and it was used in the biosensor for separating and enriching thrombin. Ethylenediamine tetraacetic acid (EDTA) was used to chelate with Ca to dissolve the hydrogel and release thrombin. A metalloporphyrinic metal-organic framework nanosheet, named as Cu-TCPP(Co) MOFs, was prepared as signal amplification strategy. Cu-TCPP(Co) MOFs/Au-ssDNA (ssDNA: single-strand DNA) was synthesized for controllable further amplification of chemiluminescent signal. The thrombin aptamer2-functionalized magnetic carbon nanotubes (MCNTs-Apt2) were used as a matrix, and Cu-TCPP(Co) MOFs/Au-ssDNA was adsorbed on the MCNTs by the complementary pairing of the partial bases between ssDNA and Apt2. Compared with ssDNA, Apt2 has a stronger interaction with thrombin. Therefore, thrombin can trigger the release of Cu-TCPP(Co) MOFs/Au-ssDNA to achieve signal amplification. Under the optimal conditions, the biosensor could detect thrombin as low as 2.178 × 10 mol/L with the range from 8.934 × 10 to 5.956 × 10 mol/L and exhibited excellent selectively. Moreover, the "signal-on" chemiluminescence biosensor showed potential application for the detection of thrombin in body fluids.
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