Construction And Application Of Bio-sensing Probes Based On Aggregation-induced Emission Materials And Long Afterglow Materials

With the progress of society and the improvement of human living standards,the health service industry based on biotechnology and life science,including bioanalysis,medical and health care,nutrition and health care,has become an important industry leading the global economic development and social progress in the 21st century.Bio-sensing research on biochemical markers can detect diseases in early stage,carry out health care and disease prevention in advance,and reduce the morbidity and mortality of the population.Nowaday,aggregation-induced emission（AIE）materials have been widely used in the field of biochemical sensing because of the excellent specificity,such as tumor marker detection,ion detection and biological imaging.However,most AIE-based probes show a"false"positive signal by fluorescence intensity due to some nonspecific and undesirable aggregation before binding to a specific receptor.Therefore,it is still a great challenge to quantify biochemical markers based on AIE probe fluorescence intensity index accurately.Lifetime is an inherent property of luminescent materials,which is rarely affected by microenvironment and measurement techniques.Therefore,probes based on lifetime signals are gradually replacing fluorescence intensity for precise quantitative detection of specific targets.However,in biological samples,AIE’s short lifetime（nanosecond level）is difficult to distinguish from biological autofluorescence lifetime（nanosecond level）.Therefore,there are operational bottlenecks in quantitative analysis by using the lifetime signal of AIE directly.In this case,we propose the idea of taking advantage of the long lifetime（millisecond level）of inorganic long afterglow nanomaterials Zn₂Ge O₄:Mn²⁺（ZGO）to avoid the interference of autofluorescence lifetime in the time domain by using time-resolution technology.However,this inorganic nanomaterial is not specific for most biomarkers.Therefore,it is difficult to accurately identify specific biomarkers.In this thesis,the research on lifetime probes based on aggregation-induced luminescent materials and long afterglow materials mainly includes the following three aspects:First,we constructed a Py TPA-ZGO lifetime probe.The probe has three lifetime signals:lifetime signal 1 from the short-lifetime AIE material Py TPA-P(τ_Pn,nanosecond level),lifetime signal 2 from the long-lifetime inorganic long afterglow material nanoparticle Zn₂Ge O₄:Mn²⁺-NH₂(τ_Zn,millisecond level),the lifetime signal 3 is a composite dual-lifetime signal(CDLSn,CDLS_n=_τPn^τZn).As the concentration of the target furin changed,the spatial distance of the donor/acceptor system composed of Zn₂Ge O₄:Mn²⁺-NH₂and Py TPA-P was corrected,and the non-radiative transition efficiency changes these three lifetime signals accordingly.These lifetime signals can be accurately quantified the concentration of target in the microenvironment.In contrast,the fluorescence intensity signal of Py TPA-P shows defectively quantitative performance.Furthermore,we find that the CDLS_nexhibits higher significant differences than the two other lifetime signals(τ_Pnandτ_Zn)thanks to its wide range between the maximum and minimum signal values and small standard deviation.Therefore,CDLS_nis further used to accurately identify cell subtypes based on the specific concentration of furin in each subtype.Lifetime criterion can realize precise quantification,and it should be a promising direction of AIE-based quantitative analysis in the future.Secondly,on the basis of the first part,we effectively extended the apparent lifetime of AIE material in the second part of the work,and constructed a novel ZPM dual-signal probe based on the apparent long lifetime signal and fluorescent signal of AIE.The probe consists of a short lifetime matrix metalloproteinase 2（MMP-2）responsive AIE material Py TPA-M and a long lifetime inorganic afterglow nanomaterial Zn₂Ge O₄:Mn²⁺-NHS（ZGO-NHS）,which acts as an energy donor to continuously deliver its own energy to the acceptor Py TPA-M,and extends the apparent lifetime of Py TPA-M from nanoseconds to milliseconds.This facilitates the detection of MMP-2without interference from the organism’s autofluorescence lifetime.With the change of target MMP-2 concentration,the apparent long litime signal and fluorescence intensity signal of Py TPA-M changed correspondingly.Therefore,based on the fluorescent signal and the apparent long lifetime signal of the ZPM dual-signal probe,we can accurately quantify the concentration of MMP-2 in He La cells in different cell cycles.The study found that the content of MMP-2 excreted in Hela cells was about twice as high as that in the cells.And MMP-2 in the cells and exocytosis was more abundantly expressed at the G1/S phase.This work will be helpful for the design and development the multi-signal probe strategy based on AIE in the future,and has important research significance in the direction of accurate quantitative analysis.Finally,based on the research on the detection function of the lifetime probe,the thrid section adds a therapeutic function to the probe by introducing a targeting peptide,and realized the integration of disease detection and treatment.Based on the apparent long lifetime signal of AIE material Py TPA-Th and the long lifetime signal of long afterglow nanomaterial Zn₂Ge O₄:Mn²⁺-NHS（ZGO-NHS）,we constructed a novel ZPTh dual-lifetime probe for imaging,detection and therapy.The probe possesses two millisecond level long lifetime signals from the thrombin-responsive AIE material Py TPA-Th(τ_Pn)and the long lifetime inorganic nanoparticle ZGO-NHS(τ_Zn).The nonradiative transition efficiency between the donor/acceptor system composed of ZGO-NHS and Py TPA-Th changed with the concentration of the thrombin,resulting in corresponding changes in the two long-lifetime signals.The ZPTh probe can detect thrombin without the interference of the autofluorescence lifetime from the organism.Simultaneously,thrombin exposes the P-selectin peptide in Py TPA-Th,enabling specific imaging of activated platelets.Due to the superior photodynamic therapy（PDT）effect of AIE material Py TPA,the early treatment of thrombosis was realized because the activated platelet apoptosis.This study provides a new idea for the rational design of AIE-based diagnosis and treatment-integrated lifetime probes.