Production of T4 antiserum and determination of its characteristics
1986
Hartono, R. (Badan Tenaga Atom Nasional, Jakarta (Indonesia). Pusat Penelitian Teknik Nuklir) | Santoso, D. | Rediatning, W. | Misyeti
One of the most important component of a radioimmunoassay kit is the antiserum, because it contains the antibodies necessary for the antigen-antibody reaction. Antisera for radioimmunoassay should meet certain requirements i.e. containing antibodies with a high specificity and avidity, a sufficient sensitivity in the desired working range and a high titre. In the present study the production of T4 antiserum was carried out by reacting L-thyroxine ethyl ester with bovine serum albumin (BSA) and the conjugate formed was used for immunization. The immunogen-adjuvant mixture was injected into rabbits, the injections being repeated at monthly intervals and the serum was collected every 2 weeks for titre determination. The results showed that a sufficiently high titre was obtained after 4 months. The quality of the antiserum produced was further evaluated by determining other antiserum characteristics like avidity and specificity. Evaluation of specificity was performed by cross reaction studies of T4 antiserum with 3,3',5-triiodo-L-thyronine (T3), 3,5-diiodo-L-thyronine (3,5-T2), 3-iodo-L-tyrosine (MIT), 3,5-diiodo-L-tyrosine (DIT), 3,3',5-triiodothyroacetic acid (TRIAC), 3,3',5,5'-tetraiodothyroacetic acid (TETRAC). The results showed that the antiserum produced has sufficient specificity for use in RIA. The antiserum avidity or K value was determined using Scatchard's plot expressing the relationship between B/F vs bound antigen. K value of 2,7 x 10 to the power of 7 1/mole was obtained from the experimental results. The sensitivity, equivalent to 1/K, which is the detection limit obtained by T4 antiserum was found to be around 3,7 x 10 to the power of -8 mole/1. It can be concluded from the experimental results that the T4 antiserum produced meet the requirement for use in RIA.
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