Evaluation The TRAb In Sera Of Human By ELISA Detecting Using Recombinent TrxTSHRc, TrxTSHRn Protein As Antigen

Autoimmune thyroid disease(AITD)is a sort of organ specificity autoimmune disease.Human thyroid stimulating hormone receptor is a primary target antigen in AITD.And plays a role in set and process of AITD.The hTSHR gene assignment at No.14 chromatin body(14q31),is a G protein-coupled large moleculer glycoprotein consisting of extracellular(TSHR-ecd),transmembrane and intramembrane domain.TSHR-ecd has multi-discontinuous binding site where the thyrotropic-stimulating hormone(TSH)and thyrotrophin receptor antibody(TRAb) can discriminate and combine.Epitopes of thyroid-stimulating antibody(TSAb) mainly lie in N terminal of TSHR-ecd(amino acid3437,40,4245,5256),while thyroid stimulating blocking antibody(TSBAb)in C terminal(amino acid 301,385,390).TSH's target organ is thyroid gland.TSH can combin with TSHR and cause a series of biochemical event by CAMP second signal system.Physiologically, the combination can regulate the normal function and growth of thyroid follicular epithelial cell.TSHR is the normal composition of the thyroid gland.But under the function of heredity and environment,the structure of TSHR changes and has the antigenicity which can cause autoimmune thyroid disease(AITD)and product autoantibodies(TRAb).The detection of TRAb is very important for the dignosis, treatment and prognostic evaluation.Objective:Using Recombinent TrxTSHRn Protein which contains TSAb antigen determinant.and TrxTSHRc Protein which contains TSBAb antigen determinant as antigen,establish enzyme linked immunosorbent assay(ELISA).for detecting the TRAb in the serum of health and thyroid disease patient.Investigate this detection's significance for the dignosis,treatment in AITD。Method:Using Recombinent TrxTSHRn Protein and TrxTSHRc Protein as antigen, determin the optimum condition,establish the indirect ELISA for detecting the TRAb in the serum of health and thyroid disease patient.Result: 1.The detection of TRAb using recombinent TrxTSHRc protein as antigen: It is the best condition for ELISA that the coating quantity of antigen is 16ng/100μl and the seru dilute 100 fold.The intra-assay coefficients of variation(CV)of positive seru and negtive seru are 3.5%,4.12%respectively.The interassay CV values are 4.52%,4.67%.The mean±standard deviation(SD)of normal value is 0.319±0.107(in adult).The positive rate of untreanted Graves' disease are 41.86%,treated Graves' disease are 7.86%,Hashimoto thyroiditis 66.01%(hypothyroidism77.78% hyperthyroidism52.94%),simple goiter are 6.98%,subacute thyroiditis are 11.1%Non-toxic nodular goiter;nontoxic nodular goiter are 8.7%thyroid adenoma are 3.1%respectively.2.The detection of TRAb using recombinent TrxTSHRn protein as antigen: It is the best condition for ELISA that the coating quantity of antigen is 100ng/100μl and the seru dilute 100 fold.The intra-assay coefficients of variation(CV)of positive seru and negtive seru are 3.77%,3.42%respectively.The interassay CV values are 4.87%,5.12%.The mean±standard deviation(SD)of normal value is 0.186±0.094(in children),0.398±0.167(in adult).The positive rate of untreanted Graves' disease are 70.31%,treated Graves' disease are 35.7%,Hashimoto thyroiditis 36.67%,simple goiter are 4.6%,subacute thyroiditis are 6.67%Non-toxic nodular goiter;nontoxic nodular goiter are 7.25%respectively.Conclusion:1.The detection of TRAb using recombinent TrxTSHRc protein as antigen is very important for the dignosis of AITD,especially in Hashimoto thyroiditis complicate hypothyroidism;2.The detection of TRAb using recombinent TrxTSHRn protein as antigen in normal:There is difference in range of normal values between children and adult,but no statistically significant between male and female;3.The detection of TRAb using recombinent TrxTSHRn protein as antigen is very important for the dignosis of AITD,especially in Graves'disease.