Quantitative Detection Of Prostate Tumor Markers Based On SERS And Fluorescence Spectroscopy

At present,it is essential that the development of cancer prevention and early diagnosis to protect the lives and health of residents,as the prevalence of cancer has increased year by year,which caused by the environmental pollution and life stress.Researchers have found that tumor markers are closely related to the development of tumors in living organisms.Therefore,some researches about tumor markers,such as the take advantage of tumor markers for early diagnosis of cancer,the constantly improvement of the sensitivity and accuracy for tumor marker detection,have become the current research hotspots.Compared with some immunologically-based tumor marker detective methods which commonly used in clinical practice,the detective method of tumor markers based on surface enhanced Raman scattering?SERS?spectroscopy and fluorescence spectroscopy technology shows higher sensitivity,more convenient operation and low-cost.As the above advantages of SERS spectroscopy and fluorescence spectroscopy technology,more and more scientists have been attracted to the research domain.However,many challenges have been faced about the application of the mentioned spectroscopy-based tumor markers detection in high-throughput,high-sensitivity,uniformity,and specific signal acquisition.Therefore,further researches on high sensitivity,high specificity and uniform signal spectroscopy have important clinical application value,trace detection of tumor markers as well.The work of this thesis mainly takes use of SERS spectroscopy and cyclic amplification fluorescence spectroscopy to quantitatively detect protein tumor markers?PSA?and gene tumor markers?miRNA-141?which related to prostate cancer.In the detection of tumor marker PSA based on SERS spectroscopy,the gold nanorod/polymethyl methacrylate?Au NRs/PMMA?substrate was prepared by Langmuir-Blodgett method and plasma surface treatment technology.Besides,the structural and optical properties of substrate were characterized.Then the specific Raman characteristic peaks of PSA have been obtained by a comprehensive analysis of the SERS spectra of male serum,female serum and alpha-fetoprotein?AFP?and human glandular kallikrein2?hK2?standard solutions.As following,the specific Raman characteristic peak of PSA has been applied to SERS spectral analysis of clinical serum samples.The experimental results show that the prepared Au NRs/PMMA substrate has high SERS enhancement and good signal reproducibility.What's more,it also presents the high specificity and sensitivity in PSA label-free detection,and the detection limit can be as low as 60 pg·mL^-1.On the other hand,in the detection of tumor marker miRNA based on cyclic amplification fluorescence spectroscopy,we designed a highly sensitive and specific detective proposal for multiple-miRNA quantitative detection by combined the fluorescence characteristics of quantum dots and the specific shear characteristics of double-strand specific nuclease?DSN?.During the experiment,the capture DNA probe has formed as the link between the quantum dots and magnetic nanoparticles with the aid of the capture DNA,then a heteroduplex DNA-miRNA hybrid structure has taken shape automatically because of the complement of DNA and miRNA,which triggered the DSN selective cleavage of the capture DNA.As the specific cleavage of DSN in the hybrid structure,the quantum dots and miRNAs could be separated from the capture DNA probe.Thereafter,a new round of hybridization and DSN specific cut will be started with the free miRNA and the unpaired DNA in the solution.Through the above cycles,the quantum dots are continuously released from the captured DNA probe,which lead to the continuously amplification of fluorescence signal.At the same time,in order to avoid the fluctuation of the serum tumor marker content?miRNA-141?caused by physiological factors,miRNA-1228 was introduced as a calibration reference,and then the quantitative detection of the multiple miRNA was performed.The results show that the specific quantitative detection of miRNA-141 and miRNA-1228 was successfully achieved by DSN-assisted circular amplification of quantum dot fluorescent signals,and the detection range was from1 fmol·L^-1 to 100 pmol·L^-1.