The Application Of Single Molecule Array In The Ultrasensitive Detection HOXC10 And CPS1 Biomarkers Of Lung Cancer

In recent years,lung cancer has become the most common malignancy and the highest mortality rate of all malignant tumors in China.The number of cases has reached 800,000 and the number of deaths is about 700,000.With the development of malignant tumors,cells,tissues or organs will release a large amount of DNA,mRNA,proteins and metabolites into the blood circulation system.The expression levels of these molecules are highly correlated with the stage of tumor development,which can be used as a biomarker for predicting the occurrence of tumor,cancer screening and clinical diagnosis.Although traditional molecular diagnostic methods,such as enzyme linked immunosorbent assay can detect common biomarkers,the vast majority of disease-related protein expression levels are too low to be detected by traditional diagnostic methods.Single-molecule array technology possesses ultra-high sensitivity and can obtain results quickly and accurately,which shows great potential in biomarker detection.HOXC10 and CPS1 are two biomarkers for diagnosis and prognostic of lung cancer that can be used for lung cancer early screening.Based on this,this thesis mainly describes the development and application of single-molecule array technology for rapidly,accurately,ultra-sensitively and quantitatively detecting HOXC10 and CPS1 biomarkers.The specific research work is as follows:1.Ultra-sensitive detection of HOXC10 biomarkers based on single-molecule array technology.By screening different HOXC10 antibodies and recombinant proteins,the most matched HOXC10 A2-A1 antigen-antibody pair was selected for ultrasensitive detection and analysis on the HD-1 Analyzer.The quantitative detection range of HOXC10 was 8-1024 pg/ml and the LOD was 1.537 pg/ml.Compared with the ELISA assay in the microplate,the sensitivity increased by about 111 times.HOXC10 had very good selectivity and could not be affected by common bio-interfering substances.The spike recovery rate of HOXC10 was between 73.4%and 88.7%,and the relative standard deviation was between 2.60%and 6.64%which showed that HOXC10 had good utility and could be directly used to detect actual samples directly.Based on these results,single-molecule array technology can quickly,ultra-sensitively and quantitatively detect biomarkers and show great potential in the field of diseases clinical diagnosis.2.Ultra-sensitive detection of CPS1 biomarkers based on single-molecule array technology.By screening different CPS1 antibodies and recombinant proteins,the most matched CPS1 A1-A2 antigen-antibody pair was selected for sensitive detection using magnetic bead-based ELISA.The quantitative detection range of CPS1 was 0.2-100 ng/ml and the LOD was 0.04 ng/ml.Moreover,CPS 1 had a very good specificity and was not affected by similar biological interferences.The spiked recovery of CPS1 was between 99.9%and 110.2%,and the relative standard deviation was between 3.72%and 5.90%which showed that CPS1 possessed good utility.Serum Validation assay indicated CPS1 exists in serum and could be used to directly detect actual samples.