Establishment Of Time-resolved Fluorescence Immunochromatographic Assay For Detection Of CAl9-9

The CA19-9 is a carbohydrate antigen which is expressed at high levels in patients with malignant tumors of the pancreatic cancer,biliary tract cancer,colon cancer,and gastric cancer.Therefore,the detection of CA19-9 in vivo can be helpful for early screening and assisting in diagnosis of malignancies.At present,there have been numerous clinical detection methods for CA19-9.With the establishment of a state hierarchical medical system,there is a crying need for a Point-of-care testing method that can be used for primary medical units.With the rapid development of diagnostic technology in vitro,fluorescence immunochromatographic assay has becoming one of the important means for POCT.Owing to this,the study intends to establish a novel fluorescence immunochromatographic method for the quantitative detection of CA19-9.Hybridoma technology was used to prepare monoclonal antibody against CA19-9,and the matched CA19-9 paired monoclonal antibody which had been screened and then was used in the establishment of fluorescence immunochromatographic assay.With the label of carboxy fluorescent microspheres and carriers of nitrocellulose membrane,suitable CA19-9 paired monoclonal antibodies was chosen to labeled and coated separately.By using the fluorescent immunochromatographic technology,the fluorescent immunochromatographic test strips for quantitative detection of serum CA19-9 content were prepared after selection of the main raw material,optimization process and optimal reaction time.According to the linear range,precision,specificity,which evaluate the fluorescence immunochromatographic assay of CA19-9 was initially established.In this study,two pairs of combinations of CA19-9 monoclonal antibody were obtained by screening.Further screening was performed by using fluorescence immunochromatography,the paired antibodies M-2E3 and M-2D8 were tested with best performance for the test strips of CA19-9.After optimization of the process,the optimal reaction time of test strips was 15 minutes.The linear equation were obtained:y=0.0069x+0.0223,the coefficient of determination was 0.9865,the linear range was 12.5-800 U/mL.The minimum detection limit of test strips was 9.74 U/mL,and the specificity was better.The Within-run and between-run CV were less than 15%.Average recovery rate was 101%.The stability of test strips was better which accelerated destruction at 37°C for 24 days.By detecting fifty clinical samples in parallel with Roche electrochemical luminescence detection kit,the results showed that there were a good correlation.The coefficient of determination was 0.9886,the regression equation y = 0.9884 x + 3.8191.In this study,CA19-9 paired monoclonal antibody was successfully prepared suitable for fluorescence immunochromatographic assay,and a fluorescence immunochromatographic assay for the quantitative detection of serum CA19-9 was established.It has a good clinical application prospect.