Development of two-step radioimmunoassay (RIA) for the measurement of free triiodothyronine in human serum based on antibody coated tubes

Journal of Radioanalytical and Nuclear Chemistry - We describe a simple, user friendly two-step radioimmunoassay (RIA) based on antibody coated tubes for the measurement of free triiodothyronine in...     

Solid phase radioimmunoassay for testosterone in human serum using antibodies coupled to magnetizable cellulose

Summary A simple user-friendly, solid phase radioimmunoassay for testosterone in human serum based on magnetic particles is described. IgG fractions precipitated from polyclonal testosterone antiserum were coupled to magnetizable cellulose particles using carbonyl diimidazole. The prepared antibody solid phase was stable for one year when stored at 4 °C. The optimized assay involves the incubation of 50 ml of testosterone standards (0.3-10 ng/ml), 100 ml of magnetizable cellulose particle coupled antibody suspension and 100 ml of ¹²⁵I-testosterone derivative for 4 hours at 37 °C. At the end of the incubation, the tubes were placed on a magnetic rack for 10 minutes after the addition of wash buffer and decanted. The sensitivity of the assay is 0.2 ng/ml. The intra-assay variation was <15% throughout the assay range. The recovery varied from 88 to 115%. On dilution of samples having high levels of testosterone, linearity ranged between 87 and 125%. Correlation coefficient of 0.978 was obtained when the developed solid phase assay was compared to the earlier established liquid phase assay.     

Standardisation of radioimmunoassay for human insulin employing magnetizable cellulose particles

We describe a convenient and flexible solid phase radioimmunoassay for human insulin employing magnetizable cellulose particles. Anti-porcine insulin antibody was covalently linked to magnetizable cellulose particles to form a stable and economical solid phase immunosorbent system. The tracer was prepared by radioiodinating insulin with ¹²⁵I using Chloramine-T oxidation method. The analytical sensitivity of assay observed was 5.5 μIU/mL. Intra-assay and inter-assay variations were found to be <12 % along with analytical recovery of 93–109 %. The developed assay can be used for the routine analysis of clinical samples. In addition, concentration of the solid phase magnetizable immunosorbent can be easily varied as per the specific requirement for research purposes.     

A method for preparation of magnetizable cellulose and its application in radioimmunoassays for T3 and T 4 1

An easy adoptable technique has been developed to prepare magnetizable cellulose by the process of wet grinding of a mixture of iron oxide (magnetite) and cellulose in a stirred ball mill. Anti rabbit IgG was covalently coupled to this magnetizable cellulose. The immunosorbent thus prepared has been used in radioimmunoassays of T₃ and T₄ for the separation of bound and free antigens. The reliability of the above immunosorbent has been validated by studying assay parameters such as non-specific binding (NSB), maximum binding (B ₀), CV etc. The single step process of wet grinding adopted here not only firmly binds magnetite and cellulose physically, but also lowers the size of the resulting magnetizable cellulose. The conventional method of preparation involves the coprecipitation of cellulose dissolved in cuprammonium hydroxide solution along with iron oxide, followed by extensive grinding and sieving of the iron oxide impregnated cellulose.