Study On The Affinity Of Serum MUC1 Antibody In Patients With Stage ? Breast Cancer

Objective:To investigate the affinity of anti-MUC1 antibodies in the serum of patients with stage IV breast cancer,and to provide basis for clarifying the binding mechanism of anti-MUC1 antibody and MUC1 antigen in stage IV breast cancer.Methods:1.Western Blot and blocking experiments: Anti-MUC1 antibody in the serum of patients with stage IV breast cancer was detected with recombinant protein 8R-MUCPT as a solid-phase antigen.Mouse anti-human MUC1 VNTR monoclonal antibody and Hepatitis B surface antigen monoclonal antibody acts as a blocking agent to block the antigen-antibody reaction;2.Indirect ELISA and inhibition experiment: Randomly selects 11 samples of serum with positive anti-MUC1 antibodies from late(IV),as well as 11 serum samples from early(non-IV)breast cancer patients.Anti-MUC1 antibodies were detected in all the serum samples using the recombinant 8R-MUCPT antigen.The same molar of 8R-MUCPT,non-8R-MUC1 recombinant protein-3a B,poly Polyarginine(Poly R)and polyhistidine(Poly H)were used as blocking agents for each sample.Then,the inhibition rates were calculated.3.Urea degeneration combined with enzyme-linked immunosorbent assay: urea was used as a denaturant to detect early cases in 61 cases and 64 in late stage of breast cancer.The anti-MUC1 antibodies in all the serum sample were detected by indirect ELISA and the affinity index were also calculated.4.Statistical analysis: SPSS 17.0 was used for the data analysis.The t-test was used for the difference analysis of the two groups of samples in the inhibition rates and affinity index.A probability value(p)of less than 0.05 was considered significant.Results:1.Western Blot blocking experiments showed that the recombinant 8R-MUCPT protein could be recognized by the natural anti-MUC1 antibody in the serum of stage IV breast cancer patients,and it could be blocked by mouse anti-human MUC1 monoclonal antibody and could not be blocked by anti-B Hepatitis surface antigen monoclonal antibody which confirms that serum anti-MUC1 antibodies can bind with MUC1 antigen in stage IV breast cancer patients;2.Indirect ELISA and inhibition experiments show that anti-MUC1 antibody in serum of stage IV breast cancer patients cannot be blocked by the 8R-MUCPT Fragments other than the MUC1 antigen,so it only specifically binds with MUC1 antigen.The inhibition rate of each sample whether in the early or late stage was exceeded 45%.There was no statistical difference in the inhibition rate between the two groups(P=0.778);3.Urea degeneration combined with enzyme-linked immunosorbent assay showed that the affinity index of anti-MUC1 antibody in serum was not significantly different between the advanced group and the early group(47.13±11.45 vs.47.53±16.10,P=0.873).Conclusion:Anti-MUC1 antibody in serum of stage IV breast cancer patients can specifically bind to MUC1 antigen;the affinity of anti-MUC1 antibody in serum of stage IV breast cancer patients is not significantly different from that in non-IV stage group.There may be insufficient binding of anti-MUC1 antibodies to serum MUC1 antigens in the stage IV breast cancer patient's serum;this insufficient binding may be related to factors other than the affinity of anti-MUC1 antibody.