Fluorescent Assay of FEN1 Activity with Nicking Enzyme-Assisted Signal Amplification Based on ZIF-8 for Imaging in Living Cells.

Flap endonuclease 1 (FEN1) participates in both DNA replication and repair to maintain the stability and integrity of the genome. As a potential tumor marker, detecting FEN1 activity could be an effective strategy for cancer diagnosis. In this work, a fluorescence assay was developed for sensitive detection of FEN1 using biomineralized metal-organic framework nanoparticles (ZIF-8 NPs) to codeliver the encapsulated proteins and DNA probes to living cells. After uptake into cells, the biomineralized ZIF-8 NPs were biodegraded to release proteins and DNA probes under an acid environment. In the presence of FEN1, the cleaved flap triggered by FEN1 hybridized with a hairpin probe to fabricate a double-stranded DNA structure which had a cleavage site of the nicking enzyme, causing the fluorophore to move away from the quencher. Assisting the nicking enzyme, an amplified fluorescence signal was obtained after several recycling. Confocal imaging indicated that this fluorescence assay could distinguish cancer cells from normal cells. Therefore, this strategy would contribute to the prediction and diagnosis in early-stage cancer.