Novel Nanostructures And Its Application In The Detection Of Fluorescent Molecules At High Concentration With Fluorescence Correlation Spectroscopy(FCS)

Fluorescence Correlation Spectroscopy is widely used for studying the dynamics and conformation change of molecules,also the interaction between molecules in and out of cells nowadays.For most of the biologically related processes,the reaction concentration are always ranging from?M-mM.Decreasing the detection volume is an effective approach to extend the up-limit of detection concentration of FCS.At present,the methods confining the detection volume include STED,TIR and various nanostructures.Compared with other methods,nanostructures are cheap and easy to fabricate,unnecessary to modify the whole FCS device.This study focused on fabricating different nanostructures and studying its applications in the FCS at high concentration.All the research contents and results are shown as follows:?1?In the first part,fabrication of large-sized Ag dimers by bridging method utilizing dithiols was studied.300 nm-diameter SiO₂@Ag composite microspheres were synthesized by reduction of[Ag?NH₃?₂]⁺ion with PVP.Through optimizing the concentration of[Ag?NH₃?₂]⁺ion,the volume of MPTMS and the kinds of reducing agents,the optical property and morphology of SiO₂@Ag were controlled.Finally,SiO₂@Ag with a plasmonic wavelength of 495 nm and covered with 1-2 nmAgNPs were obtained;Then the AuNPs derivatived with thiol function group by treating AuNPs on the slides using 1,10-decanedithiol;SiO₂@Ag dimeric structure was failed because the weak reactivity of AuNPs-SH.?2?In the seconed part,fabrication of core-satellites nanostructures utilizing“click chemistry”reaction was studied.The 70 nmAuNPs and 30nmAuNPs were derivatived with trans-cyclooctene?TCO?and1,2,4,5-tetrazine?Tz?,respectively.Along with the increase of satellites concentration,the plasmonic resonance wavelength red-shifted from 541nm to 589 nm.The relationship between apparent average numbers of satellites and concentration ratio of S vesus C fitted with a liner curve.The plamonic resonance wavelength can be tuned by changing the material kinds and sizes of cores and satellites,which indicated the feasibility of the assemble methods by“click chemistry”.?3?In the last part,the detection of high-concentration fluorescent molecules by FCS based on core-satellites nanostructures was conducted.First the quantum yield of fluorescent dye Alexa Fluor 568 was decreased to 7%when Methyl viologen dichloride hydrate was served as its quencher.With the excitation wavelength at 561 nm,the 50 nmAuNPs-30 nmAuNPs core-satellites nanostructructure was choosed as a substrate.The G?0?of FCS curve generated from 0.3?M Alexa Fluor 568 on core-satellite substrate was the same with the G?0?on a pure slide,which showed that the nanostructure lacked the ability to decrease FCS detection volume.Then the fluorescence enhancement of single core-satellite nanostructure was not observed,either.The condition of FCS instrument,the properties of core-satellites and the kinds of fluorescence dyes required more investigation.