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A coating procedure that could provide immobilization of antibodies, with increased binding capacity, that is cost effective, simple, robust, and appropriate for production scale application, is described. This coating approach of T3 antibodies to the polystyrene tubes has been systematically investigated to determine its utility for the development of coated tube Radioimmunoassay (RIA) for T3 in human serum. Further, the results obtained by the developed coating procedure are found to be comparable with those obtained by the "gold standard," the liquid phase RIA for T3. The coating procedure is completed in three major steps, each step involving an overnight incubation. The normal rabbit gamma-globulins are physically adsorbed onto the polystyrene tubes and incubated. After washing, a second antibody (goat anti-rabbit antiserum) is added and incubated. To this antigen specific antibody is added (T3 antibody produced in rabbit) and further incubated. Finally, the non-specific sites on the tubes are saturated by the blocking solution. The concentration of normal rabbit globulin, titers of second antibody and T3 antibody, and time required for coating are optimized to arrive at a suitable coating protocol. The coated tubes were evaluated for precision, reproducibility, and stability. Various parameters such as total reaction volume, incubation time and temperature, total number and volume of washings, concentration of 8-anilino-1-naphthalene sulfonic acid (ANS), and quantity of tracer per tube are optimized to arrive at a suitable standard curve. The optimized assay is validated for the quality control parameters such as intra- and inter-assay variations, recovery, and parallelism. The developed coated tubes assay had an assay range of 0.3-4.8 ng/mL with a sensitivity of 0.3 ng/mL at 90% B/B0. Batch to batch variation in coating was < 10%. The coated tubes were stable up to 1 year, which is adequate for production scale.

A composite filter pad formed by embedding activated carbon powder in fibrous matrix was applied for removal of hormones from human serum. Enhanced adsorption was demonstrated through direct comparison between the stacked filter pads and the powered carbon packed in a column both having an equal weight of carbon. In addition, the composite structure offers the advantage of eliminating the carbon fines mixed in the filtrant, avoiding the flow channeling, and providing favorable rate of adsorption. The adsorption of both thyroid hormones T3 (triiodothyronine) and T4 (thyroxine), and steroid hormone, cortisol, by the carbon filter were studied. The effect of pH on hormone adsorption by activated carbon was verified as an important environmental factor, minimum adsorption occurs at the neutral pH. It was found that T4 carrying one more iodine atom than T3 is less effectively adsorped by carbon adsorption from the serum under competitive condition due most likely to their size differences. The filter pad containing 70% activated carbon as adsorbent was fabricated in disk form and stacked in a column for processing bulk volume of biological fluid. Such a device provides an effective means for removal of hormones from biological solutions.

The performance characteristics of assays is reported for thyroxine (T4), triiodothyronine (T3), and T3-uptake (T3U) using the GAMMAFLOTM Automated Assay System. A comparison of calculated free thyroxine index (FTI) values is also presented. This automated radioimmunoassay (RIA) system utilizes a combination of continuous-flow methodology and chromatographic separation techniques. The T4 assay studied had a standard curve range of 1.5 to 24.0 microgram per dl. The intra- and inter-assay precisions were 4.3 and 5.3 percent CV, respectively, for a T4 concentration of 10.0 microgram per dl. The T3 assay had a standard curve range of 50 to 1000 ng per dl, the corresponding precisions were 7.3 and 7.1 percent CV, respectively, for a concentration of 213 ng per dl. The automated serum T4 and T3 results correlated (r = 0.966 and 0.864) with a manual radioimmunoassay procedure. Intra-assay and inter-assay precisions for a mid-range normal 30.1 percent T3U value were 6.2 percent and 4.9 percent CV, respectively. Reference range comparison of FTI by both automated and manual results correlated for 47 out of 51 (95 percent) patients compared. It is concluded that this automated system appears to offer a viable alternative to T4, T3, and T3U manual RIA techniques in terms of operational simplicity, analytical performance, and sample through-put flexibility.

The values of FT3-serum concentration (Amerlex-RIA) from 311 patients and the FT4-values (Amerlex-RIA) from 745 patients were compared with the total T3- and total T4-values with regard to their diagnostic accuracy in hypo-, eu- and hyperthyroidism. The classification of patients into these three diagnostic categories was determined by the majority of available parameters. The analysis of the predictive values demonstrated that determination of free hormones usually guarantees a safe distinction between hypo-, eu- and hyperthyroidism. Taking into account prevalence, the determination of free hormones appears to be superior to that of total hormones.

Characteristics of the T4 and T3 radioimmunoassay kits that are commercially available in the UK are described. Useful comparative data and criticism are given about many of these kits. Some unacceptable variation in accuracy is demonstrated.