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Doi:10.1016/j.cccn.2005.01.006

Clinica Chimica Acta 355 (2005) 205 – 210 Pediatric reference intervals for FSH, LH, estradiol, T3, free T3, cortisol, and growth hormone on the DPC IMMULITE 1000 Offie P. Soldina,b,c, Eve G. Hoffmanc, Michael A. Waringc, Steven J. Soldina,c,d,e,f,g,T aDivision of Endocrinology and Metabolism, Georgetown University School of Medicine, Washington D.C., United States bDivision of Cancer Genetics and Epidemiology, Georgetown University School of Medicine, Washington D.C., United States cBioanalytical Core Research Laboratory, General Clinical Research Center, Georgetown University, Washington D.C., United States dDepartment of Pharmacology, Georgetown University, Washington D.C., United States eDepartment of Pediatrics, The George Washington University School of Medicine, Washington D.C., United States fDepartment of Pathology, The George Washington University School of Medicine, Washington D.C., United States gDepartment of Laboratory Medicine, Children’s National Medical Center, Washington D.C., United States Received 15 November 2004; received in revised form 5 January 2005; accepted 7 January 2005 Background: We studied serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), triiodothyr-onine (T3), free T3 (FT3), cortisol and growth hormone (GH) concentrations in a population of pediatric patients. Thereference intervals were determined separately for females and males stratified by age groups to assess age- and sex-relateddifferences. Our objective was to obtain reference intervals for the 7 serum analytes for our pediatric population using theIMMULITE 1000 system.
Methods: Serum samples of 800 in- and out-patients, newborn to 19 years old were analyzed using the DPC IMMULITE 1000chemiluminescent immunoassay system.
Results and conclusions: We report pediatric reference intervals for FSH, LH, E2, T3, FT3, cortisol, and GH. These referenceintervals provide the basis for clinical interpretation of laboratory results using the IMMULITE 1000 system and the assessmentof child development.
D 2005 Elsevier B.V. All rights reserved.
Keywords: Free triiodothyronine; Triiodothyronine; Cortisol; Estradiol; Luteinizing hormone; Growth hormone; Gonadotropins; Newborn;Child; Pediatric T Corresponding author. Division of Endocrinology and Metab- olism and Cancer Genetics and Epidemiology, Lombardi Compre- hensive Cancer Center, LL, S-165A, Georgetown UniversityMedical Center, 3800 Reservoir Road, N.W., Washington D.C.
The gonadotropins, follicle-stimulating hormone 20057-1465, United States. Tel.: +1 202 687 4717; fax: +1 301 229 (FSH), and luteinizing hormone (LH) are synthe- E-mail address: os35@georgetown.edu (S.J. Soldin).
sized by the pituitary gland and control reproductive 0009-8981/$ - see front matter D 2005 Elsevier B.V. All rights reserved.
doi:10.1016/j.cccn.2005.01.006 O.P. Soldin et al. / Clinica Chimica Acta 355 (2005) 205–210 functions. In children, abnormalities in concentra- and out-patients as we were shifting tests to the new tions of FSH and LH can aid in the diagnosis of IMMULITE 1000 and it was essential to have age- pituitary disorders, and may be indicative of and sex-related reference intervals for our own problems in the reproductive systems of both population served in the Washington, D.C. area genders, infertility problems, early and delayed (approximately 60% African American). The refer- puberty. The ovaries, placenta, and testis synthesize ence intervals were determined separately for estradiol (E2). Estrogens are involved in develop- females and males for different age groups to assess ment and maintenance of the female phenotype, age- and sex-related differences. The Hoffmann germ cell maturation, and pregnancy. They also are approach has been used widely to evaluate reference important in longitudinal growth, nervous system intervals in the sick/hospitalized population. The maturation, bone metabolism, and endothelial validity of the reference intervals established for responsiveness in both males and females. The each of the analytes is based on analyte values from measurement of E2 is an important part of the all patients regardless of their health status, since the assessment of reproductive function in females, Hoffmann approach allows for the correction neces- including assessment of infertility, oligoamenorrhea, sary when no attempt is made to include only and menopausal status. The test also has applica- samples from individuals who were validated as tions in both men and women in osteoporosis risk being normal. A good discussion of reference assessment and monitoring of female hormone intervals and the pros and cons of different approaches to assess reference intervals have recently Cortisol (hydrocortisone) slows the inflammatory response, maintains blood pressure and cardiovas-cular function and stimulates gluconeogenesis.
Cortisol determinations are commonly used for diagnosis of Cushing’s Syndrome, congenital adre-nal hyperplasia, and adrenal insufficiency (Addi- 2.1. Patient selection and sample collection son’s disease). Increased cortisol secretion, alteredcortisol metabolism, and increased tissue sensitivity The study was conducted at Children’s National to cortisol may be related to insulin resistance, Medical Center, Washington, D.C., on routine patient serum specimens from patients age 1 day hormone (somatotropin, GH), secreted by the to 18 years accrued from January 2003 to June anterior pituitary, is essential to the processes of 2003. Specimens were sent to the laboratory after the test had been ordered by the physician. BD breakdown of triglycerides by fat cells, helps to tubes were used throughout the study with the control blood glucose, and is responsible for the majority containing serum separator. The results production of insulin-like growth factor-1 (IGF-1).
were blinded by removing all patient identifiers Low levels of GH may lead to growth retardation except age, sex, and date of specimen collection.
Approximately 50% of the samples were from minimally ill outpatients. Only a very few of the The thyroid hormone triiodothyronine (T3) can be patients would be abnormal for any one analyte. The found in the circulation—in the free (FT3) or the Hoffmann approach allows for removal of the bound form (total T3, TT3) Thyroid hormones regulate cellular metabolism and fetal neurodevelop-ment. A deficiency in thyroid hormones, thyroxine (T4) and T3 can lead to goiter and in extreme cases,if a pregnant woman is deficient the newborn can A calibrated IMMULITE 1000 system was used suffer from severe mental retardation (cretinism).
to measure analyte concentrations in serum or We studied serum FSH, LH, E2, cortisol, GH, T3, plasma. For Quality Control, 2 samples of known and FT3 concentrations in a population of 800 in- analyte concentration (DPC, Los Angeles, CA) were O.P. Soldin et al. / Clinica Chimica Acta 355 (2005) 205–210 tested daily. The patient tests were performed on laboratory mainframe and all results entered using a serum samples obtained from both hospitalized test code, the age, gender, and test result. The patients and outpatients. Blood for all sample results were collated using Stat graphics (STSC measurements was drawn by venipuncture. Prior to performing the assay, the samples were kept refri-gerated for no longer than 12 h at 2–4 8C. As we employed the Hoffmann approach, no effort wasmade to exclude subjects from the study based on The data were analyzed employing a computer adapted Hoffmann approach The data sets were separated into female and male subjects and stratified calibration range was up to 170 mU/ml and the by age. Abnormal and outlier values were truncated analytical sensitivity was 0.1 mU/ml. CVs vary from each individual age category according to the from 4.7% to 5.4% over the analytical range tested.
Hoffmann method. Generally, the top and bottom 10– 20% of the data were discarded and the central linear calibration range was up to 200 mU/ml, and the portion of the graph extrapolated. The remaining data analytical sensitivity was 0.1 mU/ml. CVs vary were either of normal Gaussian distribution or made from 3.5% to 4.6% over the analytical range tested.
to have a Gaussian distribution by calculating the Estradiol was measured in 25 Al of serum. Samples logarithm of the values to determine the 2.5th and 97.5th percentiles for each of the age groups. Percent calibration range was from 20 to 2000 pg/ml and cumulative frequency versus concentration was plot- the analytical sensitivity was 15 pg/ml. CVs vary ted to calculate the 2.5th and 97.5th percentiles. These from 6.4% to 11.8% over the analytical range were used as the final reported serum concentration tested. T3 was measured in 25 Al of serum. The calibration range was from 40 to 600 ng/dl and theanalytical sensitivity was 35 ng/dl. CVs vary from5.9% to 10.5% over the analytical range tested. Free T3 was measured in 50 Al of serum. The calibrationrange was from 1 to 40 pg/ml and the analytical The results for all the analytes are given in sensitivity was 1.0 pg/ml. CVs vary from 9.0 to 16.9 over the analytical range tested. Cortisol was broken down into the smallest possible gender and measured in 10 Al of serum. The analytical age range groups with n values sufficient to sensitivity was 0.2 Ag/dl (5.5 nmol/l). CVs vary generate reference intervals. The intervals calculated from 6.4 to 10.8 over the analytical range tested.
for the gonadotropins FSH and LH display, as ex- Growth Hormone was measured in 50 Al of serum.
The calibration range was from 40 to 600 ng/mland the analytical sensitivity was 35 ng/ml. CVsvary from 4.9% to 7.8% over the analytical range Reference intervals for follicle-stimulating hormone (FSH) was used for the quantitative measurement of FSH, LH, estradiol, T3, free T3, cortisol, and growth hormone. The IMMULITE system utilizes assay- specific, antibody or antigen-coated plastic beads as the solid phase, alkaline phosphate-labeled reagent, and a chemiluminescent enzyme substrate. Light emission is detected by a photomultiplier tube and printed reports for each sample are generated by the system computer. The data were entered into a Females and males ages newborn to 19 years old.
O.P. Soldin et al. / Clinica Chimica Acta 355 (2005) 205–210 Reference intervals for luteinizing hormone (LH) Reference intervals for triiodothyronine (T3) Females and males ages newborn to 19 years old.
Females and males ages newborn to 19 years old.
pected, a sharp increase with age (around puberty) for a To derive T3 ng/dlÂ0.0154=nmol/l.
the females and remained elevated in the older agegroups, while the LH range stayed almost constant forthe pubertal males. Sex differences were found withthe intervals for female FSHs, highest around puberty, expected, the upper limits for female estradiol are and considerably higher than the values for males (). The upper limit for FSH increases from 4.3 The intervals of T3 for both the males and to 12.0 IU/l. Our data indicate that the upper limit of females were broad during the first years of life, the range for LH increases almost threefold, from 5.0 and decreased with increase in age. In both genders, to 13.4 IU/l, between the 6- to 10-year-old and 11- to the lower limit of the range stabilized after the age 15-year-old groups, and increases further to 16.4 IU/l of 5 years, while the upper limit steadily decreased in the 15- to 19-year-old group. The reference intervals (from 320 to 202 in females and from 320 to 208 for males remained at a similar range at the 13- to 19- limit is lower than the upper limit for total T3 by a The intervals for estradiol do not display a large increase, but as in the case of the gonadotropins, the The intervals for cortisol were similar throughout upper limits of the intervals in the years of puberty, all age groups, and our results did not differentiate N11 years are noticeably higher. As would be between male and female intervals. Notice that theupper cortisol ranges are higher for sick patientsthan for healthy individuals, no doubt due to the stress of illness. We have reported this observation Reference intervals for free triiodothyronine (FT3) Females and males ages newborn to 19 years old.
Females and males ages newborn to 19 years old.
a To derive Estradiol pg/mlÂ3.67=pmol/l.
a To derive Free T3 pg/dlÂ.0154=pmol/l.
O.P. Soldin et al. / Clinica Chimica Acta 355 (2005) 205–210 A wide reference interval for growth hormone is found in the 7- to 11-year-old group, while the Reference intervals for growth hormone (GH) other age groups remained constant. Our results did not differentiate between male and female intervals.
The upper limits for growth hormone calculated in this study are much higher than those that were previously being used by the Children’s laboratory using a different kit. These new ranges are consistent with the clinical findings and data Females and males ages newborn to 19 years old.
strongly support the validity of the new intervalsfor this method ( Serum FSH concentrations are low by 6 months in TT3, and FT3 agree with earlier studies on the boys and 1–2 years in girls and increase at the onset of Abbott IMx The data of Elmlinger utilized n puberty and the development of secondary sexual values varying from 8 to 131 in the various age characteristics. The measurement of E2 is an impor- tant part of the assessment of female reproductive Despite this deficiency the data are similar to those functions, including the assessment of infertility, in this manuscript which utilizes n values falling oligoamenorrhea, and menopausal status. It is also used for monitoring ovulation induction, as well as during preparation for in vitro fertilization. The test found excellent correlation of results between these also has applications in both men and women in platforms indicating that the reference ranges can be osteoporosis risk assessment and monitoring of used for both and very likely for the IMMULITE The median concentrations of E2 and cortisol are Circulating T4 and T3 are transported mostly usually higher during the first 2 weeks after birth bound to carrier proteins, but it is FT3 that is most than thereafter. Before the onset of puberty, no active metabolically. Elevated concentrations of T3 particular sex differences were observed, and all indicate hyperthyroidism and Graves’ disease, while analyte concentrations remained relatively constant.
low concentrations indicate severe hypothyroidism. In The increase of gonadal activity in females with the normal thyroid function, total T3 (TT3) and FT3 onset of sexual maturation included an increase in concentrations show good correlation, as TT3 changes LH and FSH, which was accompanied by a strong in concentration in direct correlation with carrier increase in E2. Cortisol increased to a lesser extent proteins such that FT3 concentration remains con- during puberty. In males, the increase in hormone stant. However, in thyroid dysfunction, TT3 results concentrations was smaller. Our findings agree with may be higher while the FT3 concentrations remain earlier studies Our findings for LH, FSH, unchanged. Therefore the primary reason to selectFT3 as the analyte, in preference to TT3 test, is toimprove the accuracy for detecting hypo- and hyper-thyroidism in patients with thyroid hormone binding abnormalities that compromise the diagnostic accu- racy of total hormone measurements. It should be noted that recent tandem mass spectrometry (MS/MS) data cast doubts on the validity of TT3 measured by Females and males ages newborn to 19 years old.
a To derive Cortisol Ag/dlÂ27.6=nmol/l.
Supported by the Colaco foundation grant.
O.P. Soldin et al. / Clinica Chimica Acta 355 (2005) 205–210 [10] Soldin SJ, Murthy JN, Agarwalla PK, Ojeifo O, Chea J.
Pediatric reference ranges for creatine kinase, CKMB,Troponin I, iron, and cortisol. Clin Biochem 1999;32:77 – 80.
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nonhealthy individuals on reference interval estimation. Clin [15] Guo T, Chan M, Soldin SJ. Steroid profiles using liquid chromatography–tandem mass spectrometry with atmospheric [8] Soldin SJ, Brugnara C, Wong EC, editors. Pediatric Reference pressure photoionization source. Arch Pathol Lab Med 2004; Ranges. 4th ed. Washington (DC)7 AACC Press; 2003.
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