Preparation Of New Raman Probes And The Application In Cancer Markers Detection

In this paper, a novel method to prepare the Raman probe of branched DNA-gold nanoaggregates was proposed based on nanoparticles, DNA self-assembly techniques and surface enhanced Raman spectra (SERS). Following this, the novel Raman probe was used for simultaneous SERS detection and imaging of two biomarkers on the cancer cell surface. Then another SERS method to detect DNA methyltransferase was developed based on enzyme circulation amplification. This paper mainly includes the following aspects:1. A new method to prepare the Raman probe was explored based on Au nanoparticles and DNA self-assembly techniques. Three kinds of hairpin DNA species were designed and immobilized on the surface of gold nanoparticles (AuNPs) for the formation of DNA-gold nanoaggregates through Au-S bond. The trigger DNA can hybridize with the first hairpin DNA and a cascade of assembly steps using three hairpin DNA species was activated. After the reaction, the fabrication of branched DNA-gold nanoaggregates modified lots of the Raman dyes can be used as the Raman probe and the Raman signal can be significantly enhanced.2. A novel scheme was demonstrated for simultaneous detection and imaging of two biomarkers (MUC1 mucin and nucleolin) on the surface of a single human breast cancer cell (MCF-7) by self-assembly of branched DNA-gold nanoaggregates. Two different Raman probes were prepared to label two biomarkers. In the presence of target cancer cells, the aptamer can specifically recognize the biomarkers on the cancer cell surface leading to stem separation. The fabricated DNA-gold nanoaggregates can hybridize with the opened stem and function as Raman probes for SERS measurement and imaging. The distribution of two biomarkers can be readily visualized by SERS imaging. As compared to single target detection, simultaneous detection of two biomarkers may provide more assistance for clinical diagnosis and understanding of the magnism in cancer development.3. A new surface enhanced Raman scattering method to detect the DNA methyltransferase was developed based on the gold nanoparticles and enzyme circulation amplification. A new hairpin DNA structure contained the specific recognization of DNA mthyltransferase was designed. After the methylation, the DpnI restriction endonuclease that has the same recognition sites was added. Then a new primer DNA can be captured by the hairpin DNA modified on the magnetic microbeads. In the presence of the Raman probe and DNA Klenow polymerase, the primer DNA can be used recycled and the Raman signal can be enhanced significantly, a low detection limit of 5.0×10-6 U/μL for DNA methyltransferase was obtained.