Design, Synthesis And Biological Application Of Nano Fluorescent Probes Based On Au-Se Bonds

Gold nanoparticle(Au NP)biosensors assembled through Au-S covalent bond for imaging biomolecule have now been extensively investigated,due to their decent biocompatibility,low toxicity and unique tunable optical properties.Such nanobiotechnology of the Au NPs was first developed by Storhoff’s group and Schultz’s group,for assembling DNA onto the Au NPs via Au-S bond.Thereafter,numerous nanomaterials and biomaterials,including DNAs,RNAs,peptides and other molecules,have been reported to be assembled onto the Au NPs for bioimaging and molecular detection.However,in the process of applying the Au NP biosensors in biological systems,thiol-functionalized Au NPs encounter a competitively ligand exchange due to the abundant in vivo thiols like glutathione(GSH),resulting in detection signal distortions and rising false positive in tumor diagnosis.It is still a great challenge to solve the interference problem of biothiols encountered in the practical application of the probe.Herein,we designed two novel Au-Se bond-based nanoprobes for detecting and imaging tumor markers under high thiol environment.Such Au-Se probes with in vivo glutathione interfering resistance offer reliable detection methods in the biological and medical era.Our research work mainly include the following two parts:1.We designed a novel Au-Se bond-based nanoprobe for imaging MMP-2 in tumor cells under high thiol environment.The Au-S bond nanoprobe(Au-S-peptide-FITC)and the Au-Se bond nanoprobe(Au-Se-peptide-FITC)were firstly formed by attaching cysteine and selenocysteine to one end of the peptide,which can be specifically recognized by matrix metalloproteinases 2(MMP-2).Compared to the Au-S probes,the Au-Se probes display high thermal stability and a very good anti-interference ability towards glutathione under simulated physiological conditions.More importantly,in vitro experiments further proved that no obvious signal distortions for the tumor cells treated by the Au-Se probes was observed before and after eliminating glutathione.The high-fidelity fluorescent signal intensity be in direct proportion to the real level of MMP-2.Thus,the construction of nanosensors based on Au-Se bond with in vivo glutathione interfering resistance will provide reliable detection methods for biomolecules in the biological and medical field.2.The Au-Se bond-based nanoprobe have excellent thermal stability and strong anti-interference ability towards biothiols,so we further synthesized double-detecting Au-Se bond nano-fluorescent probes for simultaneous detection and imaging of MMP-2 and MMP-7.We assembled gold nanoprobe by coupling FITC-modified peptide and TMARA-modified onto the gold nanoparticle’s surface via Au-Se bond.The emission of FITC fluorescent dye is 520 nm which can effectively be separated with the emission 582 nm of TAMRA,so as to achieve two optically independent channels at the same time.The experimental results show that the probe with good stability can response rapidly and detect of MMP-2 and MMP-7 simultaneously in vitro and in vivo.The probe can not only avoid interference caused by biothiols but also eliminate false positive signals of the single marker under physiological conditions.This double-resistance ability allows the probe to specifically target the target molecule,enabling achieve the accuracy of diagnosis of the tumor.