A Study On Detection Of Dickkopf-1 Autoantibody In Serums Of NSCLC Patient

Background: lung cancer is the most common malignant tumor,Currently,there are many clinical methods to detect lung cancer,but these methods are not very perfect in diagnosis,treatment and prognosis of NSCLC.Tumor markers attracted a large of scholars interest because they have important function in the early diagnosis of lung cancer and possess potential applications for risk population screening. The single tumor marker for cancer detection sensitivity and specificity is not high,Combined detection of tumor markers become a trend.recently,autoantibody become a new way in tumor detection.Objective:The aims of this study is to determine what the potential clinical value of DKK-1 autoantibody was in non-small cell lung cancer.Methods: DKK-1 cDNA sequence was obtained by RT-PCR,and cloned in pGEM Teasy plasmid.the gene sequence that was correct by sequencing inserted into prokaryotic expression vector pET30a (+).The Ecoil Rosetta (DE3) bacteria was amplified and induced via IPTG after the pET30a (+)/DKK-1 translated in the E.coli, The product of expression determined the position in the Ecoil Rosetta (DE3) bacteria by cell isolation and grading,purified on a Ni-NTA His-Bind resin column, fractionated by SDS-PAGE and appraised by Western blot.At last,Plasma from 38 healthy controls and 97 non-small cell lung cancer patients were investigated for the level of DKK-1 autoantibodies by enzyme-linked immunosorbent assay (ELISA ), Serum DKK-1 autoantibodies was deteced by Western blot.in the non-small cell lung cancer patient.Results: The recombinant plasmid pET30a (+)/DKK-1 was successfully constructed by double restriction enzyme cutting and sequence-analysis.The recombinant DKK-1 protein was expressed by mean of inclusion body in the E.coil Rosetta (DE3),DKK-1 Aautoantibody of the non-small cell lung cancer patient was confirmed by Western blot.We employed the indirect ELISA method to measure the level of DKK-1 autoan- tibody in NSCLC patients and health persons serum samples.High DKK-1 autoantibody titres were found in the sera of NSCLC patients, in addition,the results showed that autoantibody immunoassay exhibited 34.0% sensitivity and 84.2% specificity.We also found that DKK -1 autoantibody level correlated with distant metastas of tumor in NSCLC patients (P < 0.05).Furthermore ,positive detection rate of NSCLC was greatly improved by detecting DKK-1 autoantibodycombined with CEA compared to other tumor markers combined with CEA in this research.Conclusion: In our research,the target protein was successfully expressed by the The recombinant plasmid pET30a (+)/DKK-1,and was applied to detect DKK-1 autoantibody in serum of NSCLC patients by the indirect ELISA method,These results may be evidence that make DKK-1 autoantibody become clinical marker for diagnosis,treatment and prognosis of NSCLC.