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Abstract
A sensitive immunoenzymometric assay (IEMA) of serum thyrotropin (hTSH) was developed using anti-hTSH rabbit polyclonal antibody and anti-hTSH in-house monoclonal antibody with a sensitivity of 0.12 mIU/L. Serum samples were incubated in ELISA wells precoated with polyclonal antibody. The hTSH bound to the wells was incubated with monoclonal antibody (detector antibody) and further with goat anti-mouse antibody–horse radish peroxidase (GAM-HRP), which obviates the need to label the detector antibody. The assay was validated by recovery, linearity, and cross-reactivity experiments with a working assay range of 0.15 to 100 mIU/L and <10% coefficient of variation (CV) for both intra- and interassay. Good correlations were obtained when compared with Immunotech hTSH IRMA (r = 0.971, n = 35). This in-house ELISA can be used as an initial screening test for thyroid dysfunction.
ACKNOWLEDGMENTS
The authors are thankful to Dr. H. D. Sarma, Head of Animal Facility and Radioisotope Laboratory, RB & HSD, BARC, for his help and support in this work. The authors are also thankful to CE, BRIT for the encouragement shown during this work.
Notes
M* – Detector monoclonal antibody; SM1, SM2, SM3 – Solid phase monoclonal antibodies; SP – Solid phase polyclonal antibody.
*For 20 assays, using different batches of reagents over a period of 32 months.