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Fluorescence resonance energy transfer-mediated immunosensor based on design and synthesis of the substrate of Amp cephalosporinase for biosensing.

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Some disadvantages, such as insufficient sensitivity and low stability of the labeled enzyme, limit further applications of this assay. In this study, a more stable enzyme, Amp cephalosporinase (AmpC), was… Click to show full abstract

Some disadvantages, such as insufficient sensitivity and low stability of the labeled enzyme, limit further applications of this assay. In this study, a more stable enzyme, Amp cephalosporinase (AmpC), was selected as the labeled enzyme, and its substrate was designed and synthesized. This substrate contained the cephalosporin ring core as the enzymatic recognition section and the structural motif of the 3-hydroxyflavone (3-HF) as the reporter molecule. AmpC can specifically catalyze the substrate and release 3-HF, which can enter the cavity of β-cyclodextrin (β-CD) on the surface of ZnS Quantum dots and form a fluorescence resonance energy transfer (FRET) signal amplification system. An AmpC-catalyzed, FRET-mediated ultrasensitive immunosensor (ACF immunosensor) for procalcitonin (PCT) was developed by combining the signal amplification system of the polystyrene microspheres and effective immune-based magnetic separation. The ACF im-munosensor has high sensitivity and specificity for the detection of PCT; its linear range is from 0.1 ng mL-1 to 70 ng mL-1, and the limit of detection can reach 0.03 ng mL-1. The spiking recoveries of PCT in human serum samples range from 98.3% to 107%, with relative standard deviations ranging from 2.14% to 12.0%. This approach was applied to detect PCT in real patient serum samples, and the results are consistent with those obtained with a commercial ELISA kit.

Keywords: fluorescence resonance; energy transfer; amp cephalosporinase; immunosensor; resonance energy; substrate

Journal Title: Analytical chemistry
Year Published: 2019

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