Academy of Diagnostics & Laboratory Medicine - Scientific Short

What are the effects of key covariates (age, sex, ethnicity) on reference values for fertility hormones in healthy children and adolescents?

Victoria Bevilacqua and Khosrow Adeli

In order to appropriately interpret laboratory test results in a clinical setting, it is crucial that reliable reference intervals be available. Establishing reference intervals in any population is a difficult and involved process; however, it is especially challenging in a pediatric population. According to the Clinical Laboratory Standards Institute (CLSI), a single reference interval study requires recruitment of at least 120 healthy individuals. Abiding by these guidelines for each analyte studied is a significant undertaking, especially considering the need for age partitioning, which is common in pediatric populations due to the effects of child development.

Given these challenges, it is not surprising that there are significant gaps in pediatric reference intervals available to clinical laboratories. However, it is nonetheless crucial that major new efforts are initiated to close these gaps. Children are not small adults and it is, therefore, important to assess the effect of development and growth on analyte concentrations in a healthy population. The CALIPER (Canadian Laboratory Initiative on Pediatric Reference Intervals) project is a national research initiative established by a Canadian team of investigators to close these critical gaps and establish a comprehensive database of age-, sex-, and ethnic origin-stratified reference values in healthy children and adolescents. The CALIPER Project has, thus far, established reference intervals for 40 biochemical markers, proteins, lipids and enzymes [1], as well as endocrine and special chemistry markers [2]. Recently, the CALIPER project also established reference intervals for 7 fertility hormones [3]. The establishment of reference intervals for these hormones of the hypothalamic-pituitary-gonadal axis is crucial as their measurement is often used in the diagnosis of pediatric endocrinopathies. Additionally, growth and development can markedly influence normal circulating concentrations of these hormones, so age-, sex-, and Tanner stage-stratified partitions were established.

A complex pattern of change in each analyte concentration from the neonatal period to adolescence was observed. As a result, many age and sex partitions were required to cover the changes in most fertility hormones over this period. The sole exception to this was prolactin, which did not demonstrate any need for sex partitions and only required 3 age partitions. All hormones studied required partitioning in the first year of life and some analytes, specifically testosterone, FSH, LH, and progesterone, also demonstrated sex differences during the early infancy period. In addition to sex differences observed in the neonatal period, as expected, several analytes also demonstrated sex differences in the teenage years. Specifically, LH, FSH, estradiol, testosterone, and progesterone were found to increase in a sex-specific manner after puberty whereas SHBG decreased.

The marked changes in analyte concentrations throughout the pediatric age range as well as the significant differences in concentrations between the sexes illustrates why covariate-stratified reference intervals are crucial in order to appropriately interpret lab test results for fertility hormones. This study takes an important step towards closing the significant gaps that currently exist in the field of pediatric reference intervals. It is important, however, to note that reference interval tables published in this new report are directly applicable to clinical laboratories using Abbott Architect reagents and would need to be further validated for use by laboratories using other immunoassay systems. The CALIPER program has recently shown transference of pediatric reference intervals for a number of biochemical markers from Abbott assays to those by other major IVD manufacturers including Beckman, Ortho, Roche, and Siemens [4]. Transference of immunoassay data may, however, be more challenging considering significant differences in assay formulation and antibodies used by different manufacturers. New efforts are underway, including new reference interval studies as well as transference studies, in order to establish full tables of pediatric reference intervals for biochemical markers and immunoassays on all major clinical laboratory instruments.


  1. Colantonio DA, Kyriakopoulou L, Chan MK, Daly CH, Brinc D, Venner AA, Pasic MD, Armbruster D, Adeli K. Closing the gaps in Pediatric Laboratory Reference Intervals: A CALIPER Database of 40 Biochemical Markers in a Healthy and Multiethnic Population of Children. Clin Chem 2012, 58(5);854-68.
  2. Bailey D, Colantonio D, Kyriakopoulou L, Cohen AH, Chan MK, Armbruster D, Adeli K. Marked Biological Variance in Endocrine and Biochemical Markers in Childhood: Establishment of Pediatric Reference Intervals Using Healthy Community Children from the CALIPER Cohort. Clin Chem 2013, 59(9):1393-405.
  3. Konforte D, Shea JL , Kyriakopoulou L, Colantonio D, Cohen AH, Shaw J, Bailey D, Chan MK, Armbruster D, Adeli K. Complex Biological Pattern of Fertility Hormones in Children and Adolescents: A Study of Healthy Children from the CALIPER Cohort and Establishment of Pediatric Reference Intervals. Clin Chem 2013, 59(8):1215-27.
  4. Estey MP, Cohen AH, Colantonio DA, Chan MK, Marvasti TB, Randell E, Delvin E, Cousineau J, Grey V, Greenway D, Meng QH, Jung B, Bhuiyan J , Seccombe D , Adeli K. CLSI-based transference of the CALIPER database of pediatric reference intervals from Abbott to Beckman, Ortho, Roche and Siemens Clinical Chemistry Assays: Direct validation using reference samples. from the CALIPER cohort, Clin Biochem 2013, 46:1197-219.

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Academy of Diagnostics & Laboratory Medicine Designation

Fellows of the Academy use the designation of FADLM. This designation is equivalent to FACB and FAACC, the previous designations used by fellows of the National Academy of Clinical Biochemistry and AACC Academy. Those groups were rebranded as Academy of Diagnostics & Laboratory Medicine in 2023.