A Clinically Useful Method for Detecting Gonadotropins in Children: Assessment of Luteinizing Hormone and Follicle-Stimulating Hormone from Urine as an Alternative to Serum by Ultrasensitive Time-Resolved Immunofluorometric Assays

Abstract

To study the feasibility of noninvasive sampling in pediatric patients, we examined the concentrations of LH and FSH in paired serum and urine samples from 65 children (age 0-15 y) with highly sensitive time-resolved immunofluorometric assays. The detection limits of the assays were 0.015 IU/L for LH and 0.018 IU/L for FSH. These sensitivity levels allowed quantification of the low prepubertal LH and FSH concentrations. The correlation between serum and urine gonadotropin values was very good (r = 0.751,~ < 0.001 for FSH; and r = 0.720,~ < 0.001 for LH), and the urine and serum concentrations were very similar. Correction of urinary gonadotropin concentrations Urine sampling has made a comeback in pediatric endocrinologic diagnostics as a result of efforts to avoid repeated, invasive blood sampling. Much interest has been focused in particular on GH in urine (1). RIA methods were not sensitive enough to measure urinary GH levels (2), but measurement of urinary GH levels became possible using more sensitive immunoassays like the immunoradiometric assay (3), sandwich enzyme immunoassay (4), and IFMA (5, 6). The situation has been similar for the gonadotropins. The presence of gonadotropins in prepubertal urine was first demonstrated in the 1960s by Fitschen and Clayton (7) and Kulin et al. (8) using bioassay and by Bagshawe et al. using RIA ( 9 ) . In 1970, FSH and L H were quantitatively measured in urine by Rifkind et al. (10) using RIA. To improve sensitivity, extraction and concentration methods were later included and bet-

Urinary beta-luteinizing hormone and beta-follicle stimulating hormone immunoenzymometric assays for population research

“…Our findings for βLH and βFSH stored long-term at 4ºC are in agreement with other studies reporting that urinary intact LH and FSH are stable without preservatives for several months at 4ºC (7,9,13). While we found no significant loss in βLH or βFSH for up to 32 weeks at −20C, other investigators report significant loss of the intact forms of these hormones when stored frozen without preservatives for 1 to 24 weeks (7,9,10,13).…”

“…While we found no significant loss in βLH or βFSH for up to 32 weeks at −20C, other investigators report significant loss of the intact forms of these hormones when stored frozen without preservatives for 1 to 24 weeks (7,9,10,13). We conclude that long term storage at refrigerated or frozen temperatures is feasible when using assays targeting the dissociated and intact forms of LH and FSH.…”

“…Most assays for LH and FSH measure the intact forms of the hormones (7)(8)(9)(10)(11)(12), and thus factors influencing dissociation of the subunits are ever present concerns. While limited storage at refrigerated temperatures (up to 6 months) does not appear to affect the stability of intact LH and FSH (7,9,13), storage at freezing or room temperature has been found to significantly reduce the amount of intact urinary LH and FSH (7,9,10,13).…”

“…While limited storage at refrigerated temperatures (up to 6 months) does not appear to affect the stability of intact LH and FSH (7,9,13), storage at freezing or room temperature has been found to significantly reduce the amount of intact urinary LH and FSH (7,9,10,13). One study, however, found no effect of up to 10 freeze-thaw cycles on urinary LH and FSH measurements (12).…”

“…Preservatives, such as glycerol, thymol, bovine serum albumin, boric acid, thimerosal, and sodium azide, or adjustment of specimen pH to neutral, are used to prevent subunit dissociation and ensure accurate measurement of intact LH and FSH (e.g., (7,9,10,(13)(14)(15)). However, preservatives have some limitations for population level research, including time and expense invested in the advance preparation of specimen collection tubes for home or field collections, potential assay interference, effects on other hormones to be measured in the same specimen, and the fact that dilution volume correction may be necessary for the final urinary result (3,13).…”