A radioimmunoassay system for LH based on the antigenic similitude of LH with HCG: methodology and applications.

A double antibody radioimmunoassay (RIA) system for LH (LH-IEP Kit) was developed using the antigenic similitude of LH with HCG. The first antibody (Ist Ab) is rabbit anti-HCG serum, initial dilution 1:200 000. The tracer is 125I-HCG (code MJ-14 Swierk Poland). The standard curve is calibrated with the reference preparation hLH-Ist-IRP 68/40 kindly offered by WHO. The IInd Ab is pig anti-rabbit IgG serum. The incubation conditions: volume-0.3 ml; time-24 hrs with Ist Ab and 24 hrs with the IInd Ab at ambient temperature. The sensitivity of the RIA system for LH is 1.5 mIU/ml. To validate our RIA system the LH was measured in the serum samples collected from 9 women during the menstrual cycles, from 2 boys during the GnRH test, from 2 amenorrheic women and from 20 children, adolescents and adults with miscellaneous pathologies. In all these samples, parallel measurements of LH and FSH were performed using DDR commercial RIA Kits-SSW. It is to be mentioned that the LH-RIA Kit-SSW is not completely homologous, the Ist Ab being rabbit serum anti-HCG. The results obtained during 4 menstrual cycles, in which the LH peak is observed around the mid point of the interval are: follicular phase 17.92 +/- 5.58 mIU/ml (means +/- SD) with LH-IEP-Kit and 5.51 +/- 2.77 mIU/ml with the LH-SSW-Kit, peak: 26.07 +/- 22.13 mIU/ml and 9.13 +/- 5.60 mIU/ml respectively; luteal phase: 12.55 +/- 5.46 mIU/ml and 3.4 +/- 2.38 mIU/ml, respectively. The LH values observed by the two kits through all 9 menstrual cycles are well correlated ("r" values in the range 0.7-0.9) but high discrepancies were observed in the remaining 3 cycles ("r" between 0.07 and 0.6). These discrepancies as well as those observed in some adolescents with genetic anomalies and in a patient at climacterium are suggesting that the two LH-RIA systems measure not only a common molecular area but also different areas of the LH circulating molecules.