Determination of a common genetic variant of luteinizing hormone using DNA hybridization and immunoassays

OBJECTIVE. An immunologically anomalous form of LH, due to two point mutations in codons 8 and 15 of the LHβ gene, has previously been described. LH status, i.e. the discrimination between wild-type (WT) and variant (V) LH, is usually determined by immuno assays, which can be unreliable at low serum concentrations of LH. A DNA hybridization assay was therefore developed to score the LH genotype in all subjects, independent of their serum LH concentrations. To evaluate the performance of the hybridization method, and to expand our observations of the worldwide occurrence of the V-LH, we determined its frequency in additional populations. To confirm the connection between the anomalous immunoreactivity and the V-LHβ gene, we also sequenced the LHβ subunit gene of a homozygous person. DESIGN. According to the ratio of two immunoassays, one detecting only WT-LH and the other detecting equally WT and V-LH, individuals can be classified as homozygotes for the V-LHβ allele, heterozygotes or WT. DNA samples from persons with known LH status, according to the immunoassays, were used for the development and evaluation of a new allele-specific DNA hybridization assay. This assay, and PCR and restriction fragment length polymorphism analysis, were used to determine the frequency of the V-LHβ allele in DNA samples obtained from eight populations. PATIENTS. Ambulatory adult men and women, apparently healthy and with no endocrine disorders. RESULTS. The LH genotyping by immunoassays and by the new hybridization method gave identical results with all samples analysed (n=25). The V-LHβ subunit was observed to always have the two point mutations, and to be identical with the ones previously reported. The V-LHβ carrier frequency in the DNA samples collected from various populations varied between 0 and 53.5%. CONCLUSIONS. The immunoassay technique and the hybridization assay can be used as alternatives to determine the LH status. A great variation in carrier frequency of the V-LHβ allele is observed in different populations.