A simple immunoradiometric assay for human follicle stimulating hormone (hFSH) was developed using a pair of monoclonal antibodies obtained from commercial sources. The system developed makes use of a capture antibody covalently coupled to magnetisable cellulose, which is a more economical and stable immunosorbent as compared to the other solid phases. The detector antibody is labeled with 1251 using the chloramine-T oxidation method and purified by gel filtration. After initial cross-matching of the capture and detector antibodies, various assay parameters have been optimised. This assay does not show any significant cross reactivity with homologous hormones. A number of serum samples from men and women from reproductive age group was screened and compared with another commercially available kit (r = 0.98). Sensitivity of the assay is 1.4 mlU/ml, interassay variation is <5% and intraassay variation around 15%. The assay is reproducible and sensitive enough for regular estimation of serum hFSH and is relatively inexpensive.
This paper describes the use of avidin-biotin interaction as an affinity system, wherein avidin immobilized magnetizable particles (cellulose) are used as a common separation system in immunoradiometric assay (IRMA) for hormones of the human reproductive system, human follicle stimulating hormone (FSH), and luteinizing hormone (LH). Biotinylated probe was prepared by biotinylation of specific monoclonal antibody for respective antigen using the caproyl derivative of biotin N-hydroxysuccinimide. The detector antibody for the respective antigen was radiolabelled with 125I by a chloramine-T oxidation method and purified by gel filtration. In the IRMA procedure, standard/sample, respective biotinylated, and radiolabelled antibody as a single reagent, and avidin solid phase were added simultaneously to the assay tubes. After incubation for 3 h with shaking, the bound complex was quantitated for its radioactivity associated with the common avidin solid phase. Results showed that the developed assay protocol is applicable to IRMA of FSH and LH with good precision (intra and inter assay CV less than 8% and 11%, respectively), good assay range (0-200 mIU/mL) and analytical recovery (87-110%). The assay could detect 0.5 mIU/mL and 0.9 mIU/mL of FSH and LH, respectively, and showed good correlation with commercially available kits (FSH y = 0.98x + 0.21 and LH y = 0.99x + 0.18).
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