Aggregation-induced Luminescence Enhancement Of DNA-protected Silver Nanoclusters And Its Application In Biological Detection

DNA-protected silver nanoclusters(AgNCs)have the advantages of good light stability,good biocompatibility and adjustable emission range,which are attracting more and more attention in the application of diagnosis and treatment.The combination of DNA-protected AgNCs with a recognizing DNA aptamer has further expanded its use in biological assays and bioimaging.In recent years,it has been found that fluorescent metal nanoclusters have aggregation-induced luminescence enhancement(AILE)characteristics,which have a very significant effect on improving the luminescence properties of metal nanoclusters,thus enhancing their application prospect in detection and imaging.Therefore,we synthesize AgNCs by designing reasonable DNA sequences.We want to regulate the luminescence properties of DNAprotected AgNCs and expand their applications in biomolecule detection based on AILE effects via appropriate regulatory factors.In this paper,AgNCs-dsDNA,which are protected by double-stranded DNA,were designed and synthesized.The interactions between them and specific proteins were studied.The process was monitored by luminescence spectroscopy to examine whether the AgNCs had AILE characteristics and reveale relative molecular mechanisms.Based on these research ideas,we mainly carried out two parts of studis:1.The interaction of AgNCs-dsDNA with plasmodium falciparum Lactate dehydrogenase(pfLDH)induced luminescence enhancement and combined with specific DNA aptamers to achieve trace detection of pfLDH.The main biological function of pfLDH is the conversion of pyruvate to lactic acid,which is an important biomarker of malaria induced by Plasmodium falciparum.At the concetration of 1 μM,AgNCs-dsDNA has a two-step luminescence response to pfLDH so that it can achieve a wide range detection of pfLDH(≤ 1500 nM).The limit of detetion(LOD)was reduced by reducing the concentration of AgNCs-dsDNA to 0.10 μM.The calculated LOD reached 0.2 nM(7.4 pg/μL),which fell within the serum concentration range of clinical patients(3-15 pg/μL).Moreover,this method was successfully used to detect pfLDH in fetal bovine serum,demonstrating its potential application prospect in malaria clinical diagnosis.In addition,we also studied the selectivity,anti-interference ability and response to pH.Finally,by adding the aptamer 2008 s,the luminescence enhancement of AgNCs-dsDNA and pfLDH complex system was partially quenched so that we can distinguish pfLDH from plasmodium vivax lactate dehydrogenase(pvLDH)and human lactate dehydrogenase(hLDH).We further studied the the luminescence enhancement machnism of AgNCs-dsDNA and pfLDH.(1)The luminescence enhancement of AgNCs-dsDNA and pfLDH was partial quenched by the aptamer 2008 s.From that we could get that one of the binding site between AgNCs-dsDNA and pfLDH was the same as that of the aptamer.In the two-step luminescence enhancement response,the second step luminescence intensity significant enhancement is associated with the site of 2008 s.The first step of slow enhancement may be related to other sites pending further study.(2)The interaction between pfLDH and AgNCs-dsDNA limited the vibration and rotation of ligand DNA,reducing the energy loss caused by non-radiation,which induced the luminescence enhancement.(3)We further studied the effect of G-rich on the enhancement process by comparing the luminescence enhancement ratio of AgNCsssDNA with pfLDH,and we deduced that the electron-rich G-rich enhanced ligandmetal charge transport(LMCT)or ligand-metal-metal charge transport(LMMCT),which caused luminescence enhancement.2.The interaction of AgNCs-dsDNA with BSA could induce a maximum luminescence enhancement about 5 folds and blue shift.We attributed this enhancement to nonspecific electrostatic interactions between BSA and ligand double-stranded DNA that produced aggregates that limited the vibrations and rotations of ligand DNA and reduced energy loss caused by nonradiative decay.We could observe loose sheet structure by transmission electron microscopy(TEM).Furthermore,the optical properties of AgNCs-dsDNA were further altered by adding trypsin to the blends of AgNCs-dsDNA and BSA resulting in a maximum luminescence enhancement about 30 times,mainly due to the further aggregation of AgNCs-dsDNA induced by BSA hydrolyzate.Through TEM,we could observe the formation of more dense nanospheres with diameters greater than 50 nm.It is also explained by the UV-Vis spectra and the time-resolved luminescence spectra.After the formation of the nanospheres,the shortlived disappearance indicates that,in essence,the luminescence enhancement is due to providing a better protected environment after the aggregation of AgNCs-dsDNA that separated AgNCs-dsDNA from water to reduce the role of water quenching and after aggregation,limiting the intramolecular ligand DNA vibration and rotation,reducing energy loss caused by the non-radiation that improved the proportion of radiation transitions.Therefore,we have successfully regulated the optical properties of AgNCsdsDNA through the BSA and trypsin or chymotrypsin.The method can expand to regulate the optical properties of other metal nanoclusters,via the selection of appropriate protein and enzyme system.