Development of Competitive Enzyme
- Author:
Myoung Lyeh PARK
;
Youn Kyu KIM
;
Chang Kyu KIM
;
Jong Bae KIM
- Publication Type:Original Article
- Keywords:
Triiodothyronine(T3);
anti-T antibody
- MeSH:
Enzyme-Linked Immunosorbent Assay;
Glutaral;
Hyperthyroidism;
Immune Sera;
Immunization;
Linear Models;
Metabolism;
Methods;
Sensitivity and Specificity;
Staphylococcal Protein A;
Thyroid Gland;
Triiodothyronine
- From:Journal of Korean Society of Endocrinology
1996;11(4):447-454
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
Background: Triiodothyronine(T3) is a hormone secreted from thyroid gland which exerts a stimulating effect on metabolism. The disorder of thyroid system brings about several serious diseases like hypothymidism or hyperthyroidism. Therefore, the determination of T in blood is very important on monitoring thyroid function. Methods: Rabbit anti-T3 antibody was generated by immunization of T-BSA as an immunogen and purified hom antisera using Affi-gel protein A kit. The titer and specificity of purified antibody were characterized. To detect T3, competitive ELISA was performed using anti-T3 antibody and T3-HRP conjugate which was synthesized by glutaraldehyde method. The sensitivity and precision assay wer~e deterrnined and compared with that of RIA. Results: The titer of purified anti-T3 antibody was about 1:100 and the optimal dilution of T3- HRP conjugate was 1:1000. When the standard curve was constructed by ELISA, its sensitivity was about 0.5ng/ml. The eoefficient variations of intra- and inter-assay were 4.9~9.3% and 7.5~13.8%, respectively. The results obtained by ELISA and RIA correlated well with each other(n =50, r= 0.97), The linear regression equation was y= 1.09*0.08(P<0.01). Conclusion: We successfully developed a method for the measurement of T3 on ELISA which was based on competitive reaction between antigen(T3) and enzyme labeled antigen(T3-HRP). These results demonstrated that competitive ELISA is a convenient, fast, reproducible and aecurate method for the determinstion of T in serum and can be used as practical alternative to RIA.
