Preparation Of Organic Fluorescent Probes For Biosensing And Bioimaging Research

Organic aggregation-induced emission fluorescent probe is an important branch in the research area of fluorescent probe.It has great potential for chemical sensing,bioimaging and disease diagnosis and turns out to be an intersectant research field between chemistry and biology,which induces increasing attention by scientists.This kind of fluorescent probe is a hot research field,and it is still a challenge to develop simple small molecular probes with great optical property and recognition ability.We mainly designed a series of novel aggregation-induced emission fluorescent probes,which perform high luminious efficiency.By connecting different recognition groups to these probes,or introducing other fluorescent quenching materials to the sensing system,we successfully realize biosensing or bioimaging towards target substances with rather low background noise,high sensitivity and selectivity.1.Synthesis of 9,10-distyrylanthracene Derivatives of Small Molecular Fluorescent Probe and Heavy Metal Ions AnalysisWe designed a novel 9,10-distyrylanthracene?DSA?derivative with thymine groups called DSA-T2,and studied its aggregation-induced emission?AIE?property.DSA-T2 almost has no emission in the good solvent of acetonitrile.When adding mercury ions into solution,the coordination between DSA-T2 and mercury ions will take place,which largely restricts the intramolecular single bond rotation of DSA-T2 and reduces non-radiative transition,resulting in strong fluorescence signal in solution.While other metal ions have no coordination towards DSA-T2,and the solution still has no emission.This small molecular fluorescent probe is easily prepared with high sensitivity and selectivity.Besides,another small molecular fluorescent probe named DSAI with quaternary ammonium salt groups is used for silver ion detection.DSAI can aggregate on the DNA chain through electrostatic interaction and hydrophobic interaction,which leads to strong fluorescence of DSAI.We choose an aptamer?a single-strand DNA?as the recognizer of silver ion,which can induce the aggregation of DSAI to form a strong emission.The addition of silver ions into the solution will shift the conformation of the aptamer to achieve a hairpin structure,which further enhances the emission of DSAI.After adding nuclease S1 into solution,this hairpin structure will not be hydrolyzed and the emission will stay still.While other metal ions have no influence for aptamer conformation,thus aptamer will be hydrolyzed into fragments in the presence of nuclease S1,which cannot induce the aggregation of DSAI,resulting in no emission of the solution.This fluorescent probe has good selectivity and high sensitivity with the excellent fluorescent linear relationship.It realizes the turn-on and label-free detections for heavy metal ions.2.Biosensors Based on 9,10-distyrylanthracene Derivative of Fluorescent Probe and Water-soluble Carbon NanotubesWe have found that DSAI can aggregate on DNA chain to generate fluorescence.Rely on that,we developed a biosensor for ATP analysis with an AIE molecule as the fluorescent probe,a ss-DNA aptamer as the recognition part,and water-soluble carbon nanotubes as the selective part with the fluorescent quenching effect.Water-soluble carbon nanotubes can interact with ss-DNA aptamer through hydrogen bonds,thus the aptamer will be absorbed on the surface of water-soluble carbon nanotubes and the fluorescence of aggregated DSAI molecules will be quenched by water-soluble carbon nanotubes.When adding ATP into the solution,the specific binding between the aptamer and target ATP is quite strong and makes the aptamer released from the water-soluble carbon nanotubes.As a consequence,the AIE-active probes,also released from water-soluble carbon nanotubes,recover to give fluorescence and present a strong emission.It is a simple,turn-on and labelfree biosensor for specific detection of ATP.Besides,we studied the interactions between DSAI and double-strand DNA and found that embedded interaction played a key role.Based on that fact,we design an optical platform for SNP detection by using DSAI as the fluorescent probe and water-soluble carbon nanotubes as the selective platform via the fluorescent quenching effect.The embedded interactions between DSAI and complete complementary DNA?wild DNA?are so strong that the embedded DSAI molecules cannot be dragged from wild DNA by the water-soluble carbon nanotubes and the fluorescence stays still.Whereas the mutate DNA has single mutate base,the embedded interactions between mutate DNA and DSAI are relatively weak,therefore some DSAI molecules combining with mutate DNA through electrostatic interactions and hydrophobic interactions will attach on the water-soluble carbon nanotubes,leading to the partial fluorescence quenching in the solution.In this way,the single base mutate DNA can be distinguished from wild DNA.This optical biosensor shows great potential with label-free,simple and universal properties.3.Preparation of Tetraphenyl Ethylene Bioaptasensor for Cytokine BioimagingWe designed an aptamer modified tetraphenyl ethylene?TPE?derivative biosensor for interferon-gamma?IFN-gamma?detection in live cells.The TPE molecule is connected on the end of aptamer to increase the solubility of this fluorescent sensor.This bioaptasensor will interact with IFN-gamma to emit red fluorescence,which is benefit for bioimaging.We studied the labelling effect for normal BV2 cells,and there was no labelled cells due to the few content of IFN-gamma in BV2 cells.In contrast,if a small amount of IFNgamma was injected into BV2 cells for incubation,there were labelled cells with red emission.To further testify the labelling accuracy of this bioaptasensor towards IFNgamma,T cells named PBMC with relatively high concentration of IFN-gamma was chosen for labelling.As a result,we could see the obvious red emission in the cytoplasm of cells.We also used unmodified TPE molecule for comparative experiment,and the T cells were not labelled.Therefore,this bioaptasensor presents great selectivity and sensitivity,which is an advantage to label the low level of cytokines in live cells.4.Preparation of Near-infrared Magnetic Fluorescent Nanoparticles for Cytokine BioimagingNear-infrared magnetic fluorescent nanoparticles named Fe₃O₄/DSACN@PLGA/anti-VEGF NPs are produced with biodegradation property.Poly?lactic-coglycolic acid??PLGA?is used as the shell structure,oleic acid coated magnetic Fe₃O₄ and oil-soluble DSA-CN molecules are loaded inside the shell,Vascular endothelial growth factor antibody?anti-VEGF?is linked outside the shell,which has a specific combination with VEGF-A.The fluorescence of aggregated DSA-CN cannot be completely quenched by Fe₃O₄,thus the nanoparticle has the functions of fluorescence imaging and magnetic resonance imaging.Cancer cells has rich content of VEGF-A,and the antibody modified nanoparticles are available to label cancer cells by recognizing VEGF-A.With the heat given off by Fe₃O₄ after exposed under near-infrared light,some cancer cells will be killed.This nanoparticle also has good biodegradation due to the degradable PLGA shell.Two cancer cells are chosen for labelling,one is HCT116 cell and another is Raji cell.The result shows that the labelling efficiency of Raji cells is higher than that of HCT116 cells,because Raji cells contain more content of VEGF-A.On the contrary,BV2 cells contain few VEGFA and present no emission during labelling.It testifies the good selectivity of this nanoparticle towards VEGF-A.In conclusion,these near-infrared magnetic fluorescent nanoparticles have good selectivity and sensitivity,easy preparation,low biotoxicity,great biocompatibility and biodegradation,which are suitable for bioimaging.