Quantitative Analysis Of Complex Systems By Combining Spectral Sensing Technologies And Novel Calibration Strategies

Both fluorescence spectroscopy and surface-enhanced Raman spectroscopy are powerful analytical techniques.However,when being applied to the quantification of the analytes of interest in complex systems,their quantitative results are generally unsatisfactory due to detrimental effects caused by variations in factors such as impurity interference,test conditions,and samples’physical properties,etc.Therefore,the correction and elimination of the detrimental effects of these interfering factors is crutial for improving accuracy of both fluorescence and SERS techniques for the quantative analysis of complex systems.Recently,our research group developed the spectral shape deformation quantitative theory（SSD）and generalized standard addition multivariate calibration strategy based on probe technology(GMSA_probe)which can mitigate the detrimental effects of various interfering factors on the quantitative results of spectral techniques.In this thesis,effects have been made to integrate these two novel calibration strategies with both SERS and fluorescence sensing techniques for quantitative analysis of complex systems.The details of this thesis are as follows:In Chapter 2,SSD was combined with SERS technique to mitigate the effects caused by variations in the physical properties of SERS enhanced substrates on the quantitative results of SERS assays.Experimental results demonstrated that the proposed method could successfully realize rapid and accurate quantification of5-fluorocytosine（5-FC）in pills,serum and plasma samples.In Chapter 3,a simple but effective fluorescence sensing method for DNA detection was proposed by integrating exonuclease III assisted signal amplification and probe-based target fishing strategy.The proposed fluorescence sensing method in combination with GSAM_probe was applied to the quantification of Mn SOD DAN in lysate samples of Hela,Jurkat and Ramos cells.Experimental results revealed that the detection limit of the proposed method for Mn SOD DNA was 0.42 p M.And its recovery rates were in the range of 90.2%to 111.2%.These results fully demonstrated that GSAM_probecould effectively correct the detrimental effects of both background interferences and matrix effects of complex systems,and help realizeaccurate quantification of the target DNA in complex biological samples.In Chapter 4,efforts have been made to further explore the application potential of the method developed in Chapter 3.The detective sensitivity of the method was further enhanced through increasing the number of fishing times,and satisfied the requirement of the detection of mi RNA in cells.Experimental results revealed that the detection limit of the enhanced method for mi RNA-155 in cells of KH-2 and BRSA-2B was about 36 f M.The recovery rates were in the range of 96.2%to 105%,which were in good consistent with the corresponding results obtained by q RT-PCR.These results further demonstrated the generality of the proposed method and its robustness to fluorescence interferences presented in complex biological samples.