New Method Of Polymeric Nanoparticles-Assisted Cascade Amplification And Its Application In Molecular Diagnosis

Aiming at the key problems of sensitivity and operability for molecular detection, this paper combines the complex nanoparticles with glucose to construct a new method of signal amplification. The composite nanoparticles with high surface area, make the surface local enrichment of high concentrations of initiator and thus rapid triggered redox polymerization generates a signal amplification, which achieves the uncorrected visual detection of biomolecules and improves the sensitivity of the detection. In the presence of glucose molecules, GOx consumes oxygen in the air to produce hydrogen peroxide which can form redox system with vitamin C on the surface of the composite nanoparticles to initiate monomer in situ polymerization and achieve visual amplification effect. Composite nanoparticles and GOx chain amplification achieved operation in the air and improve the detection sensitivity of the multiple purposes. Specific work of this paper is as follows:1. Construction of composite nanoparticles:Amphilic block polymer PAAA-b-PS and biotin-PEG4ooo-b-PS were synthesized by RAFT polymerization, and the behavior of co-assembly was researched by DLS and TEM. Polystyrene is a hydrophobic core, and the hydrophilic shell is composed of PAAA with reduction and biotinylated PEG. Using the characteristic of nanoparticles with large specific surface area, composite nanoparticle hydrophilic shell enriched with reducing vitamin C and vitamin H, which can form a redox pair with hydrogen peroxide and as a molecular recognition of the connection link, respectively.2. Herein we established a H2O2-NP redox polymerization-assisted signal amplification system for the detection of DNA and partial p53 gene. The detection limits were 100 fM (0.3 amol) and 10 pM (30 amol), respectively. The time of surface polymerization was 30 min.3. The composite nanoparticles were combined with GOx, and the polymerization conditions such as GOx and glucose were optimized by near infrared spectroscopy. Under the condition of GOx 1.0 mg/mL and glucose 250 mg/mL, the detection sensitivity of the model DNA was 10 fM (0.15 amol) and the p53 was 5 pM (15 amol). And the time of signal amplification was 40 min.