Contruction Of Highly Sensitive Rare Earth Upconversion Fluorescence Nanosensors And Their Applications

In recent years,lanthanide-doped upconversion nanoparticles?UCNPs?which are able to emitting higher-energy visible light under near-infrared light excitation?typically 980 nm?via a two-photon or multiphoton mechanism have aroused considerable attention.Compared with quantum dots?QDs?and fluorescent dyes,UCNPs possess several outstanding features such as low toxicity,tunable emission,greater tissue penetration,good chemical and physical stability,and reducing excitation light scattering.More importantly,the near-infrared excitation light of UCNPs is not absorbed by biological samples in the detection process,which could eliminate the background autofluorescence,thus providing an enlarged signal-to-noise ratio and improved sensitivities.Thanks to these advantages,UCNPs have high application value in biosensing.Typically,most analytical applications are based on the fluorescence resonance energy transfer?FRET?principle to quench the fluorescence of UCNPs.However,the conventional absorber is hard to quench the fluorescence of UCNPs,which limits the FRET efficiency and reduces the sensitivity of the FRET-based analytical system.Therefore,it is of great significance to further improve FRET efficiency or explore new strategies to construct highly sensitive and highly selective nanosensors.Based on the above considerations,this thesis synthesized excellent UCNPs by hydrothermal method and combined different signal amplification strategies to construct several fast,reliable,highly sensitive and highly selective nanosensors for the detection of target analytes?ion and biomolecules?.The main research contents are as follows:1.A novel label-free fluorescence nanosensor based on FRET between silver triangular nanoplates?STNPs?and UCNPs was developed for ultrasensitive detection of protamine and trypsin.In this assay,negatively charged STNPs are introduced to quench the fluorescence of the positively charged UCNPs by electrostatic interaction.When protamine is added to the system,STNPs will first interact with it and then detach from the surface of UNCPs.This leads to the recovery of the fluorescence of UCNPs.Trypsin could catalyze the hydrolysis of protamine and effectively quench the fluorescence recovered by protamine.By measuring the changes of the fluorescence of UCNPs,the concentrations of protamine and trypsin were determined.Under the optimized conditions,the linear response range was obtained from 10 to500 ng/mL,5 to 80 ng/mL and with the low detection limit of 3.1 ng/mL and 1.8ng/mL for protamine and trypsin,respectively.Meanwhile,the nanosensor shows good selectivity,sensitivity and can be successfully applied to detection of protamine and trypsin in serum samples.2.A nanosensor based on inner filter effect?IFE?between UCNPs and squaric acid?SQA?-iron?III?was designed for highly sensitive and selective detection of glucose.In this assay,GOx-catalyzed oxidization of glucose produces gluconic acid and H₂O₂.The latter can catalytically oxidize Fe²⁺to Fe³⁺which can rapidly?<1 min?coordinate with the SQA to produce SQA-Fe³⁺.The absorption band of SQA-Fe³⁺largely covered the emission band of UCNPs,resulting the fluorescence emission of UCNPs was effectively quenched.Under the optimal condition,the fluorescence quenching efficiency was linearly related to the glucose concentration in the ranges of7-110?M?R²=0.987?and 110-340?M?R²=0.997?,respectively.The developed method has been further applied to monitor glucose levels in human serum with satisfactory results.3.A novel ultrasensitive upconversion fluorescence nanosensor was developed for the detection of Cu²⁺that initially integrates analyte-triggered cyclic autocatalytic amplification with L-cysteine–mediated?L-cys–mediated?aggregation of label-free gold nanoparticles?AuNPs?.The principle of the strategy is that the AuNPs can quench the fluorescence of UCNPs and the addition of L-cys can induce the aggregation of AuNPs,resulting that the fluorescence of UCNPs recovery.Upon addition of Cu²⁺,the Cu²⁺can catalyze oxidize L-cys to L-cystine and the Cu²⁺is reduced to Cu⁺,the Cu⁺can be oxidized circulatory to Cu²⁺by dissolved O₂,which catalyze and recycle the whole reaction.Thus,the aggregation of AuNPs is inhibited and the fluorescence recovered by L-cys is effectively quenched.Under the optimal condition,the fluorescence quenching efficiency shows a good linear response to Cu²⁺concentrations in the ranges of 0.4-40 nM with a detection limit of 0.16 nM,which is5 orders of magnitude lower than the U.S.Environmental Protection Agency limit for Cu²⁺in drinking water?20?M?.The method has been further applied to monitor Cu²⁺levels in real samples and the results of the method are consistent well with that gained by atomic absorption spectroscopy?AAS?.4.A novel upconversion fluorescent and colorimetric dual-readout assay was designed for H₂O₂ as well as H₂O₂-related analytes via enzyme-controlled cyclic signal amplification strategy.As a proof of application demonstration,the highly sensitive detection of choline and acetylcholine chloride?ACh?was performed.This sensing strategy involves the reaction of ACh with acetylcholinesterase?AChE?to generate choline which is followed catalytically oxidized by choline oxidase?ChOx?to produce H₂O₂.The generated H₂O₂ can transform Fe²⁺to Fe³⁺and free hydroxyl radical?·OH?.The as-produced Fe³⁺and·OH can simultaneously oxidize o-phenylenediamine?OPD?to 2,3-diaminophennazine?OPDox?and the Fe³⁺was reduced to Fe²⁺.The Fe²⁺produced from the redox processes between Fe³⁺and OPD could be cycle used to generate OPDox that can significantly quench UCNPs fluorescence based on IFE.On the other hand,colorimetric signal can be visualized by the naked eye and applied to directly recognize the concentration of choline and ACh.The phenomenon is further designed as a colorimetric logic gate.Moreover,we also detected H₂O₂,glucose,lactate and uric acid?UA?with the higher sensitivity by utilizing the assay strategy,which revealed the extensive applications of the proposed sensing strategy in biomedical analysis.5.A novel ultrasensitive upconversion intense red fluorescence and colorimetric dual-readout strategy was developed for the determination of alkaline phosphatase?ALP?that initially integrates enzyme-triggered cascade signals amplification?ECSAm?with rapid fusing reaction of label-free STNPs.The sensing strategy involves four aspects.Firstly,STNPs with an absorption band at?610 nm was synthesized,which can effectively quench UCNPs fluorescence at 670 nm.Secondly,when in the presence of ALP,L-ascorbic acid 2-phosphate?AAP?was transformed into ascorbic acid?AA?which can react with KIO₃ to produce I₂.Thirdly,the procreant I₂ can quickly adsorb onto the surface of STNPs,then etch STNPs and the I₂is reduced to I^-.Fourthly,the generated I^-can further accelerate the fusion of STNPs by adsorption effect,which helps achieve ECSAm,allowing the quantitative evaluation of ALP with a satisfying detection limit of 0.035 mU/mL.In addition,the developed method has been further applied to the detection of ALP in human serum with satisfactory results.These results indicate that the dual-readout assay with a well-defined response mechanism shows good prospects in physiological and pathological studies.