Local preparation and evaluation of double-antibody liquid phase radioimmunoassay system for detection of human testosterone

Abstract EN

Preparation, evaluation and optimization of testosterone radioimmunoassay (RIA) system using liquid phase double antibody is considered to be the main objective.

Three primary components were prepared and characterized to obtain valid and accurate system.

These components were polyclonal testosterone antibody, the 125I-testosterone tracer and set of testosterone standards.

The production of polyclonal testosterone antibody was undertaken by immunizing two groups of females white New-Zealand rabbits with testosterone-3-(O-carboxy methyl oxime): BSA as immunogen through primary immunization and five boosters.

Both R1 and R4 gave anti-serum has a high immunoreactivity.

The preparation of 125I-testosteronetracer was carried out using three different conjugates (testosterone-3-TME, testosterone-3-histamine and testosterone-3-BSA) by electrophilic substitution mechanism using chloramine-T as oxidizing agent.

Tracers were characterized in terms of radiochemical yield %, radiochemical purity %, specific activity and immunoreactivity.

A set of testosterone standards were prepared using highly purified testosterone antigen.

Optimization and validation tests of the local liquid phase RIA system were carried out.

In conclusion, the results showed that, the local testosterone RIA system is sensitive, specific and accurate with significant cost reduction in comparison with commertial kit and extended use of the method for routine investigation of variety of diseases especially hypogonadism and associated male infertility.