Establishment Of Chemiluminescent Immunoassay Methods For Three Tumor Markers

Ovarian malignancy is one of the common gynecological malignancies.The incidence rate is second only to cervical cancer and uterine body cancer,but the mortality rate of ovarian cancer is higher than that of other gynecologic cancers,which poses a serious threat to women’s lives.Because the early symptoms are not typical,70%of patients are diagnosed with tumor cells have spread to other organs of the body,only about 30%of ovarian cancer patients are found that the tumor confined to the ovary in the operation,so early diagnosis of ovarian cancer has become a significant problem.The use of a single tumor marker for early diagnosis of ovarian cancer often presents specificity and low sensitivity.Carbohydrate antigen CA125 and human epididymis protein HE4 are important markers of ovarian cancer.CEA,a tumor marker of the broad-spectrum tumor,is expressed in many kinds of tumors.Clinically,multiple tumor markers are often used jointly for the detection to assist the early diagnosis of ovarian cancer.This project established CEA plate chemiluminescence immunoassay method,and CA125,HE4 magnetic particle chemiluminescence immunoassay method.The experiments optimized the important parameters of the reagents,achieved the best detection results,achieved a quantitative detection of three tumor markers,and reassessed the linearity,sensitivity,reproducibility and specificity and other properties of the detection method.The results showed that the linear range of the CEA plate chemiluminescence detection method which is established in this study was 0-32 ng/ml,and the sensitivity was 0.146 ng/ml.The precision experiments showed that the intra-and inter-assay CV were 6.8%-8.5%and 7.7%-11.7%.There was no cross-reaction with several common serum tumor markers in it.The test results of commercial ELISA kit are of good correlation,with the correlation coefficient is greater than 0.99.This study successfully produced anti-CA125 and anti-HE4immunomagnetic microspheres as well as established CA125,HE4 magnetic particle chemiluminescence detection method.The results showed that CA125 and HE4 are of good correlation within the linear range of 0-1200 U/ml and 0-1000 PM/L respectively,with R~2≥0.99;the intra-assay variations of the two magnetic particle chemiluminescence detection methods were all less than 6%;results of the cross-reaction experiment showed that there was no cross-reaction between this method and the common serum tumor markers;degree of the detection sensitivity of CA125 and HE4 were 0.25 U/ml and 0.5 PM/L.The spike recovery experiments showed that the matrix effect of the detection method was low.The methodological comparison results showed that the method established in this thesis is highly related to the detection results of commercial ELISA kits.The three tumor markers constructed in this study provide the basis for applying multiple markers jointly in diagnosing ovarian cancer.