Normative Values of High-Sensitivity Cardiac Troponin T and N-Terminal pro-B-Type Natriuretic Peptide in Children and Adolescents: A Study from the CALIPER Cohort.

BACKGROUND: Cardiac troponin (cTn) and N-terminal pro-B-type natriuretic peptide (NT-proBNP) are increasingly used clinically to evaluate and prognosticate acute myocardial infarction and heart failure, respectively. Pediatric reference intervals and cut-offs have not been established for Roche's Elecsys Troponin T hs (high sensitive) assay. Although pediatric reference intervals exist for NT-proBNP, cut-off values do not exist. In this study, we report reference intervals and 99th percentile cut-offs in a large, healthy Canadian pediatric population using the CALIPER cohort. METHODS: Blood samples from 484 healthy children and adolescents between 0 and <19 years old were recruited from hospital outpatient clinics and community settings. Serum samples were analyzed using Roche's Cobas e411 and evaluated for high-sensitivity cTnT (hs-cTnT) and NT-proBNP concentrations. 95% reference intervals and 99th percentile cut-off values were established. RESULTS: Three hs-cTnT age partitions were established (0 to <6 months, 6 months to <1 year, and 1 to <19 years) with highest concentrations observed in children under 1 year. Two NT-proBNP age partitions were established (0 to <1 year, and 1 to <19 years), also with higher concentrations in infants under 1 year of age. For each of these age partitions, the 99th percentile cut-off, 95% reference interval, and proportion of detectable concentrations were determined. CONCLUSIONS: This is the first study to examine hs-cTnT and NT-proBNP reference values together in a healthy pediatric cohort without other clinical indications. We present 99th percentile cut-offs, which will allow clinicians to appropriately evaluate cardiovascular disease in children and adolescents.

CLSI-based transference and verification of CALIPER pediatric reference intervals for 29 Ortho VITROS 5600 chemistry assays.

BACKGROUND: Evidence-based reference intervals (RIs) are essential to accurately interpret pediatric laboratory test results. To fill gaps in pediatric RIs, the Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) project developed an age- and sex-specific pediatric RI database based on healthy pediatric subjects. Originally established for Abbott ARCHITECT assays, CALIPER RIs were transferred to assays on Beckman, Roche, Siemens, and Ortho analytical platforms. This study provides transferred reference intervals for 29 biochemical assays for the Ortho VITROS 5600 Chemistry System (Ortho). METHODS: Based on Clinical Laboratory Standards Institute (CLSI) guidelines, a method comparison analysis was performed by measuring approximately 200 patient serum samples using Abbott and Ortho assays. The equation of the line of best fit was calculated and the appropriateness of the linear model was assessed. This equation was used to transfer RIs from Abbott to Ortho assays. Transferred RIs were verified using 84 healthy pediatric serum samples from the CALIPER cohort. RESULTS: RIs for most chemistry analytes successfully transferred from Abbott to Ortho assays. Calcium and CO2 did not meet statistical criteria for transference (r2<0.70). Of the 32 transferred reference intervals, 29 successfully verified with approximately 90% of results from reference samples falling within transferred confidence limits. Transferred RIs for total bilirubin, magnesium, and LDH did not meet verification criteria and are not reported. CONCLUSIONS: This study broadens the utility of the CALIPER pediatric RI database to laboratories using Ortho VITROS 5600 biochemical assays. Clinical laboratories should verify CALIPER reference intervals for their specific analytical platform and local population as recommended by CLSI.

Pediatric reference intervals for 29 Ortho VITROS 5600 immunoassays using the CALIPER cohort of healthy children and adolescents.

BACKGROUND: Accurate reference intervals (RIs) based on a healthy pediatric population are essential for pediatric test result interpretation. The CALIPER project has recruited a large healthy cohort and completed a series of a priori studies to address gaps in pediatric RIs. As immunoassays from different manufacturers for endocrine and special chemistry markers are not standardized and show marked intermethod differences, direct RI studies are needed for each major analytical platform. Here, we report age- and sex-specific pediatric RIs for 29 immunoassays on the Ortho Clinical Diagnostics (Ortho) VITROS® 5600 analyzer. METHODS: Health information and blood samples were collected from healthy pediatric subjects. Using the Ortho VITROS 5600 Integrated System MicroWell Technology, 29 biomarkers were measured. Analyte concentrations were partitioned by age and sex according to the Harris and Boyd method. After removing outliers, age- and sex-specific RIs and corresponding 90% confidence intervals were calculated according to CLSI guidelines. RESULTS: All analytes required age partitioning except ß-human chorionic gonadotropin (ß-hCG), cancer antigen 15-3 (CA15-3), rubella immunoglobulin G (rubella IgG), and vitamin D. Several analytes including estradiol, progesterone, testosterone, follicle-stimulating hormone (FSH), luteinizing hormone (LH), free triiodothyronine (FT3), total triiodothyronine (TT3), total thyroxine (TT4), thyroid uptake, ferritin, intact parathyroid hormone (iPTH), total prostate-specific antigen (tPSA), free prostate-specific antigen (fPSA), cancer antigen 125 (CA125), creatine kinase MB (CK-MB), and myoglobin showed sex differences, observed mostly with the onset of puberty. CONCLUSIONS: Complex reference value trends were observed across the pediatric age range for several biomarkers examined on Ortho VITROS immunoassays. The availability of VITROS immunoassay RIs will enable accurate laboratory test interpretation and diagnosis for the pediatric population. As recommended by the CLSI EP28-A3c guidelines, implementation of these RIs should be validated for each laboratory's local pediatric population.

Pediatric Reference Intervals for Transferrin Saturation in the CALIPER Cohort of Healthy Children and Adolescents.

BACKGROUND: Transferrin saturation reference intervals specific for age and sex have not been previously reported for the pediatric population. The reference values for transferrin saturation have been previously reported to be lower in children compared to adults, caused by a combination of low serum iron and high serum transferrin levels in children, warranting specific reference intervals. Here we use the original iron and transferrin data from the CALIPER cohort to establish age- and sex-specific pediatric reference intervals for transferrin saturation. METHODS: Iron and transferrin concentrations were measured in serum samples from the CALIPER cohort of healthy children and adolescents on the Abbott Architect c8000. Transferrin saturation was subsequently calculated and statistically relevant age- and sex-partitions were determined. After removing outliers, age- and sex-specific reference intervals with corresponding 90% confidence intervals were calculated using CLSI C28-A3 guidelines. RESULTS: Transferrin saturation required 3 separate age partitions, with an additional sex partition for 14-<19 year olds. Transferrin saturation was more variable during the first year of life, evident by a wider reference interval, which subsequently narrowed at one year until adolescence. Upon adolescence, a sex difference was apparent with females having lower percent transferrin saturation than males. CONCLUSIONS: Age- and sex-specific pediatric reference intervals for transferrin saturation were established based on a large cohort of healthy pediatric subjects. Transference studies suggest that these intervals established using Abbott assays are comparable to those on Beckman, Ortho, Roche, and Siemens assays. Individual laboratories should however verify these reference intervals for their individual instrument and local population as per CLSI guidelines.

Pediatric reference value distributions and covariate-stratified reference intervals for 29 endocrine and special chemistry biomarkers on the Beckman Coulter Immunoassay Systems: a CALIPER study of healthy community children.

BACKGROUND: The CALIPER program is a national research initiative aimed at closing the gaps in pediatric reference intervals. CALIPER previously reported reference intervals for endocrine and special chemistry markers on Abbott immunoassays. We now report new pediatric reference intervals for immunoassays on the Beckman Coulter Immunoassay Systems and assess platform-specific differences in reference values. METHODS: A total of 711 healthy children and adolescents from birth to <19 years of age were recruited from the community. Serum samples were collected for measurement of 29 biomarkers on the Beckman Coulter Immunoassay Systems. Statistically relevant age and/or gender-based partitions were determined, outliers removed, and reference intervals calculated in accordance with Clinical and Laboratory Standards Institute (CLSI) EP28-A3c guidelines. RESULTS: Complex profiles were observed for all 29 analytes, necessitating unique age and/or sex-specific partitions. Overall, changes in analyte concentrations observed over the course of development were similar to trends previously reported, and are consistent with biochemical and physiological changes that occur during childhood. Marked differences were observed for some assays including progesterone, luteinizing hormone and follicle-stimulating hormone where reference intervals were higher than those reported on Abbott immunoassays and parathyroid hormone where intervals were lower. CONCLUSIONS: This study highlights the importance of determining reference intervals specific for each analytical platform. The CALIPER Pediatric Reference Interval database will enable accurate diagnosis and laboratory assessment of children monitored by Beckman Coulter Immunoassay Systems in health care institutions worldwide. These reference intervals must however be validated by individual labs for the local pediatric population as recommended by CLSI.

Transference of CALIPER pediatric reference intervals to biochemical assays on the Roche cobas 6000 and the Roche Modular P.

OBJECTIVES: The Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) has recently established pediatric age- and sex-specific reference intervals for over 85 biochemical markers on the Abbott Architect system. Previously, CALIPER reference intervals for several biochemical markers were successfully transferred from Abbott assays to Roche, Beckman, Ortho, and Siemens assays. This study further broadens the CALIPER database by performing transference and verification for 52 biochemical assays on the Roche cobas 6000 and the Roche Modular P. DESIGN AND METHODS: Using CLSI C28-A3 and EP9-A2 guidelines, transference of the CALIPER reference intervals was attempted for 16 assays on the Roche cobas 6000 and 36 on the Modular P. Calculated reference intervals were further verified using 100 healthy CALIPER samples. RESULTS: Most assays showed strong correlation between assay systems and were transferable from Abbott to the Roche cobas 6000 (81%) and the Modular P (86%). Bicarbonate and magnesium were not transferable on either system and calcium and prealbumin were not transferable to the Modular P. Of the transferable analytes, 62% and 61% were verified on the cobas 6000 and the Modular P, respectively. CONCLUSIONS: This study extends the utility of the CALIPER database to two additional analytical systems, which facilitates the broad application of CALIPER reference intervals at pediatric centers utilizing Roche biochemical assays. Transference studies across different analytical platforms can later be collectively analyzed in an attempt to develop common reference intervals across all clinical chemistry instruments to harmonize laboratory test interpretation in diagnosis and monitoring of pediatric disease.

Complex reference value distributions and partitioned reference intervals across the pediatric age range for 14 specialized biochemical markers in the CALIPER cohort of healthy community children and adolescents.

BACKGROUND: The CALIPER program has previously reported a comprehensive database of pediatric reference intervals for 63 biochemical and immunochemical markers. Here, covariate-stratified reference intervals were determined for a number of special assays not previously reported. METHODS: A total of 1917 healthy children and adolescents were recruited and serum concentrations of 14 biochemical markers were measured using the Abbott Architect ci4100 system. Age and gender partitions were statistically determined, outliers removed and reference intervals calculated using CSLI C28-A3 guidelines. RESULTS: Many analytes showed dynamic changes in concentration requiring at least 3 age partitions. Unique intervals were required within the first year of life for: pancreatic amylase, C-peptide, ceruloplasmin, insulin, ß-2-microglobulin, cystatin C, dehydroepiandrosterone sulfate (DHEA-S), and α-1-glycoprotein. Cholinesterase, cholinesterase-dibucaine number, and immunoglobulin E required only 2 age partitions and α-1-antitrypsin required only one. Anti-CCP and anti-TPO levels were below the detection limit of the assay. Some analytes including insulin and DHEA-S required additional gender partitions for specific age groups. CONCLUSIONS: Complex profiles were observed for endocrine and special chemistry markers, requiring establishment of age- and gender-specific reference intervals. These updated reference intervals will allow improved laboratory assessment of pediatric patients but should be validated for each analytical platform and local population as recommended by CLSI.

CLSI-based transference of the CALIPER database of pediatric reference intervals to Beckman Coulter DxC biochemical assays.

BACKGROUND: The CALIPER program has established a comprehensive database of age- and sex-stratified pediatric reference intervals for over 85 common biochemical markers, largely using the Abbott ARCHITECT assays. To allow a broader application of the CALIPER database, we examined transference to 36 Beckman Coulter Synchron Unicel DxC800 assays, based on the CLSI C28-A3/EP9-A3 guidelines. METHODS: Patient sample comparisons were performed for 36 biochemical assays using 200 serum specimens obtained from pediatric patients on the Abbott ARCHITECT ci8200 and the Beckman Coulter DxC800. For each analyte, R(2) values were calculated to assess the quality of correlation between the platforms. Statistical criteria used to assess transferability included a) regression analysis to create the equation of the line of best fit, b) standardized residual, c) Bland-Altman, and d) quantile-quantile plots. Transferred reference intervals were further verified by analyzing serum samples from 100 healthy children from the CALIPER cohort on the Beckman Coulter system. RESULTS: The reference intervals for most of the assessed analytes were transferable to Beckman Coulter assays (31 out of 36 studied) and the newly calculated reference intervals were verified through analysis of CALIPER reference samples (28 out of 31). Eighteen assays demonstrated excellent correlation (R(2)≥0.95), and 13 assays showed strong correlation (0.77≤R2≤0.94). CONCLUSION: The current study allowed successful transference of a large number of biochemical markers from the CALIPER database to assays on the Beckman Coulter DxC800 platform. Transference should facilitate broader application of CALIPER reference intervals at pediatric centers using DxC biochemical assays.

CLSI-based transference of CALIPER pediatric reference intervals to Beckman Coulter AU biochemical assays.

OBJECTIVE: The CALIPER program has established a comprehensive database of pediatric reference intervals using largely the Abbott ARCHITECT biochemical assays. To expand clinical application of CALIPER reference standards, the present study is aimed at transferring CALIPER reference intervals from the Abbott ARCHITECT to Beckman Coulter AU assays. DESIGN AND METHODS: Transference of CALIPER reference intervals was performed based on the CLSI guidelines C28-A3 and EP9-A2. The new reference intervals were directly verified using up to 100 reference samples from the healthy CALIPER cohort. RESULTS: We found a strong correlation between Abbott ARCHITECT and Beckman Coulter AU biochemical assays, allowing the transference of the vast majority (94%; 30 out of 32 assays) of CALIPER reference intervals previously established using Abbott assays. Transferred reference intervals were, in general, similar to previously published CALIPER reference intervals, with some exceptions. Most of the transferred reference intervals were sex-specific and were verified using healthy reference samples from the CALIPER biobank based on CLSI criteria. It is important to note that the comparisons performed between the Abbott and Beckman Coulter assays make no assumptions as to assay accuracy or which system is more correct/accurate. CONCLUSION: The majority of CALIPER reference intervals were transferrable to Beckman Coulter AU assays, allowing the establishment of a new database of pediatric reference intervals. This further expands the utility of the CALIPER database to clinical laboratories using the AU assays; however, each laboratory should validate these intervals for their analytical platform and local population as recommended by the CLSI.

Dynamic biological changes in metabolic disease biomarkers in childhood and adolescence: A CALIPER study of healthy community children.

BACKGROUND: Understanding age- and sex-specific biological changes in metabolic disease biomarkers is essential for their appropriate utilization in management of children with inborn errors of metabolism (IEM). The CALIPER program aimed to establish pediatric reference values in healthy community children for common metabolic biomarkers and determine the effects of key covariates including age and sex across the pediatric age. METHODS: A cohort of 500 healthy children and adolescents from birth to 19years were initially recruited to establish pediatric reference intervals according to the CLSI C28-A3 guidelines. Serum samples were used to measure 37 amino acids by ultra-performance liquid chromatography, 32 acylcarnitines, as well as free and total carnitine by tandem mass spectrometry, and ß-hydroxybutyrate and free fatty acids using the Vitros 5.1 chemistry analyzer. P ediatric reference intervals were calculated using non-parametric statistics and partitioned based on age- and sex-distributions. RESULTS: Approximately 80% of all analytes required 2 to 4 age-dependent partitions, with over 50% of amino acids and over 70% of acylcarnitines exhibiting significant physiological changes during the neonatal period. Also, 21% of all analytes required partitioning during puberty and adolescence, half of which produced sex-specific distributions. CONCLUSIONS: A comprehensive reference interval database for metabolic disease biomarkers established in this study will improve detection of IEMs by providing appropriate age- and sex-related information in the pediatric population. It will also aid newborn screening programs and guide the management of patients with known metabolic diseases, especially pubertal and adolescent boys and girls that display sex-specific concentrations.

Clinical decision limits for interpretation of direct bilirubin--a CALIPER study of healthy multiethnic children and case report reviews.

OBJECTIVE: Measurement of total and direct bilirubin is routinely performed for the differential diagnosis of hyperbilirubinemias. The diagnostic efficiency of a test is dependent on the chosen clinical decision limit. This study is designed to address the clinical decision limits for direct bilirubin. DESIGN AND METHODS: Routine laboratory method was used to measure total and direct bilirubin in children up to the age of 18years. Case study data and serum from a group of healthy children were analyzed and statistical exercise was performed to establish decision limits. RESULTS: The reference interval for total bilirubin was 1-12µmol/L and for direct bilirubin 1-9µmol/L with the median direct bilirubin of 3µmol/L. In 17% of children with non-pathological jaundice, median total bilirubin was 173µmol/L, median direct bilirubin was 8µmol/L and median direct bilirubin percent was 49%. From birth direct bilirubin percentage decreased until total bilirubin was 41µmol/L, then it remained at ≤10%. Albumin increased with age, and was on average 2.4g/L higher when measured using bromocresol-green compared with bromocresol-purple. An increased amount of direct bilirubin was observed when albumin (detected using the bromocresol-purple method) was >35g/L. CONCLUSIONS: Direct bilirubin concentration of ≥10µmol/L should be used to consider the presence of conjugated hyperbilirubinemia provided that total bilirubin is also above the reference interval. A high direct bilirubin percentage is unlikely to offer any clinical value when total bilirubin is not increased. It is, however, a useful diagnostic tool when there is a persistence of hyperbilirubinemia or when total bilirubin increases during times of stress with direct bilirubin >10%.

Pediatric population reference value distributions for cancer biomarkers and covariate-stratified reference intervals in the CALIPER cohort.

BACKGROUND: Cancer biomarkers are commonly used in pediatrics to monitor cancer progression, recurrence, and prognosis, but pediatric reference value distributions have not been well established for these markers. The Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) sought to develop a pediatric database of covariate-stratified reference value distributions for 11 key circulating tumor markers, including those used in assessment of patients with childhood or adult cancers. METHODS: Healthy community children from birth to 18 years of age were recruited to participate in the CALIPER project with informed parental consent. We analyzed serum samples from 400-700 children (depending on the analyte in question) on the Abbott Architect ci4100 and established reference intervals for α-fetoprotein (AFP), antithyroglobulin (anti-Tg), human epididymis protein 4 (HE4), cancer antigen 125 (CA125), CA15-3, CA19-9, progastrin-releasing peptide (proGRP), carcinoembryonic antigen (CEA), squamous cell carcinoma antigen (SCC), and total and free prostate specific antigen (PSA) according to CLSI C28-A3 statistical guidelines. RESULTS: We observed significant fluctuations in biomarker concentrations by age and/or sex in 10 of 11 biomarkers investigated. Age partitioning was required for CA153, CA125, CA19-9, CEA, SCC, proGRP, total and free PSA, HE4, and AFP, whereas sex partitioning was also required for CA125, CA19-9, and total and free PSA. CONCLUSIONS: This CALIPER study established a database of childhood reference intervals for 11 tumor biomarkers and revealed dramatic fluctuations in tumor marker concentrations between boys and girls and throughout childhood. In addition, important differences between the adult and pediatric population were observed, further highlighting the need for pediatric-specific reference intervals.

Pediatric within-day biological variation and quality specifications for 38 biochemical markers in the CALIPER cohort.

BACKGROUND: Studies of biological variation provide insight into the physiological changes that occur within and between study participants. Values obtained from such investigations are important for patient monitoring and for establishing quality specifications. In this study we evaluated the short-term biological variation of 38 chemistry, lipid, enzyme, and protein analytes in a pediatric population, assessed the effect of age partitions on interindividual variation, and compared the findings to adult values. METHODS: Four plasma samples each were obtained within 8 h from 29 healthy children (45% males), age 4-18 years. Samples were stored at -80 °C and analyzed in 3 batches, with samples from 9-10 study participants per batch. Within-person and between-person biological variation values were established using nested ANOVA after exclusion of outliers by use of the Tukey outlier test. Analytical quality specifications were established with the Fraser method. RESULTS: Biological variation coefficients and analytical goals were established for 38 analytes. Age partitioning was required for 6 analytes. Biological variation characteristics of 14 assays (37%) were distinct from adult values found in the Westgard database on biological variation. Biological variation characteristics were established for 2 previously unreported analytes, unconjugated bilirubin and soluble transferrin receptor. CONCLUSIONS: This study is the first to examine biological variation and to establish analytical quality specifications on the basis of biological variation for common assays in a pediatric population. These results provide insight into pediatric physiology, are of use for reference change value calculations, clarify the appropriateness of reference interval use, and aid in the development of quality management strategies specific to pediatric laboratories.

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.

BACKGROUND: Pediatric endocrinopathies are commonly diagnosed and monitored by measuring hormones of the hypothalamic-pituitary-gonadal axis. Because growth and development can markedly influence normal circulating concentrations of fertility hormones, accurate reference intervals established on the basis of a healthy, nonhospitalized pediatric population and that reflect age-, gender-, and pubertal stage-specific changes are essential for test result interpretation. METHODS: Healthy children and adolescents (n = 1234) were recruited from a multiethnic population as part of the CALIPER study. After written informed parental consent was obtained, participants filled out a questionnaire including demographic and pubertal development information (assessed by self-reported Tanner stage) and provided a blood sample. We measured 7 fertility hormones including estradiol, testosterone (second generation), progesterone, sex hormone-binding globulin, prolactin, follicle-stimulating hormone, and luteinizing hormone by use of the Abbott Architect i2000 analyzer. We then used these data to calculate age-, gender-, and Tanner stage-specific reference intervals according to Clinical Laboratory Standards Institute C28-A3 guidelines. RESULTS: We observed a complex pattern of change in each analyte concentration from the neonatal period to adolescence. Consequently, many age and sex partitions were required to cover the changes in most fertility hormones over this period. An exception to this was prolactin, for which no sex partition and only 3 age partitions were necessary. CONCLUSIONS: This comprehensive database of pediatric reference intervals for fertility hormones will be of global benefit and should lead to improved diagnosis of pediatric endocrinopathies. The new database will need to be validated in local populations and for other immunoassay platforms as recommended by the Clinical Laboratory Standards Institute.

Marked biological variance in endocrine and biochemical markers in childhood: establishment of pediatric reference intervals using healthy community children from the CALIPER cohort.

BACKGROUND: Reference intervals are indispensable in evaluating laboratory test results; however, appropriately partitioned pediatric reference values are not readily available. The Canadian Laboratory Initiative for Pediatric Reference Intervals (CALIPER) program is aimed at establishing the influence of age, sex, ethnicity, and body mass index on biochemical markers and developing a comprehensive database of pediatric reference intervals using an a posteriori approach. METHODS: A total of 1482 samples were collected from ethnically diverse healthy children ages 2 days to 18 years and analyzed on the Abbott ARCHITECT i2000. Following the CLSI C28-A3 guidelines, age- and sex-specific partitioning was determined for each analyte. Nonparametric and robust methods were used to establish the 2.5th and 97.5th percentiles for the reference intervals as well as the 90% CIs. RESULTS: New pediatric reference intervals were generated for 14 biomarkers, including α-fetoprotein, cobalamin (vitamin B12), folate, homocysteine, ferritin, cortisol, troponin I, 25(OH)-vitamin D [25(OH)D], intact parathyroid hormone (iPTH), thyroid-stimulating hormone, total thyroxine (TT4), total triiodothyronine (TT3), free thyroxine (FT4), and free triiodothyronine. The influence of ethnicity on reference values was also examined, and statistically significant differences were found between ethnic groups for FT4, TT3, TT4, cobalamin, ferritin, iPTH, and 25(OH)D. CONCLUSIONS: This study establishes comprehensive pediatric reference intervals for several common endocrine and immunochemical biomarkers obtained in a large cohort of healthy children. The new database will be of global benefit, ensuring appropriate interpretation of pediatric disease biomarkers, but will need further validation for specific immunoassay platforms and in local populations as recommended by the CLSI.

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.

OBJECTIVES: The CALIPER program recently established a comprehensive database of age- and sex-stratified pediatric reference intervals for 40 biochemical markers. However, this database was only directly applicable for Abbott ARCHITECT assays. We therefore sought to expand the scope of this database to biochemical assays from other major manufacturers, allowing for a much wider application of the CALIPER database. DESIGN AND METHODS: Based on CLSI C28-A3 and EP9-A2 guidelines, CALIPER reference intervals were transferred (using specific statistical criteria) to assays performed on four other commonly used clinical chemistry platforms including Beckman Coulter DxC800, Ortho Vitros 5600, Roche Cobas 6000, and Siemens Vista 1500. The resulting reference intervals were subjected to a thorough validation using 100 reference specimens (healthy community children and adolescents) from the CALIPER bio-bank, and all testing centers participated in an external quality assessment (EQA) evaluation. RESULTS: In general, the transferred pediatric reference intervals were similar to those established in our previous study. However, assay-specific differences in reference limits were observed for many analytes, and in some instances were considerable. The results of the EQA evaluation generally mimicked the similarities and differences in reference limits among the five manufacturers' assays. In addition, the majority of transferred reference intervals were validated through the analysis of CALIPER reference samples. CONCLUSIONS: This study greatly extends the utility of the CALIPER reference interval database which is now directly applicable for assays performed on five major analytical platforms in clinical use, and should permit the worldwide application of CALIPER pediatric reference intervals.

Long-term stability of biochemical markers in pediatric serum specimens stored at -80 °C: a CALIPER Substudy.

OBJECTIVES: Pediatric serum samples collected from healthy children in the CALIPER (Canadian Laboratory Initiative in Pediatric Reference Interval) project are stored at -80 °C for various periods of time. This study aimed to determine the stability of chemistry, protein, and hormone analytes under these conditions. DESIGN AND METHODS: Serum samples collected from children of 0-18 years of age attending outpatient clinics were pooled into a single pool or into age-group specific pools. Following baseline measurement, each pool was aliquoted and kept frozen at -80 °C until analysis. Samples were analyzed for 57 biochemical markers at monthly intervals over a 10-13 month period and each aliquot was subject to one freeze-thaw cycle before analysis. The analysis was performed on VITROS Chemistry System, COBAS INTEGRA 400 Plus and IMMULITE 2500. Values obtained at monthly intervals were compared to baseline measurements and examined for trends over time. RESULTS: A majority of analytes measured in this study showed no significant time-dependent change relative to baseline or trend over time after up to 13 months of storage. PTH showed up to 27.2% decline after 10 months of storage with most of the decline evident after 2 months. Most analytes showed variability over time, which is thought to reflect assay variability rather than changes in analyte stability. CONCLUSIONS: The study shows stability for a majority of analytes stored in serum at -80 °C after up to 13 months of storage. Samples do not require immediate testing for reference interval determination for the selected analytes with possible exception of PTH.

Closing the gaps in pediatric laboratory reference intervals: a CALIPER database of 40 biochemical markers in a healthy and multiethnic population of children.

BACKGROUND: Pediatric healthcare is critically dependent on the availability of accurate and precise laboratory biomarkers of pediatric disease, and on the availability of reference intervals to allow appropriate clinical interpretation. The development and growth of children profoundly influence normal circulating concentrations of biochemical markers and thus the respective reference intervals. There are currently substantial gaps in our knowledge of the influences of age, sex, and ethnicity on reference intervals. We report a comprehensive covariate-stratified reference interval database established from a healthy, nonhospitalized, and multiethnic pediatric population. METHODS: Healthy children and adolescents (n = 2188, newborn to 18 years of age) were recruited from a multiethnic population with informed parental consent and were assessed from completed questionnaires and according to defined exclusion criteria. Whole-blood samples were collected for establishing age- and sex-stratified reference intervals for 40 serum biochemical markers (serum chemistry, enzymes, lipids, proteins) on the Abbott ARCHITECT c8000 analyzer. RESULTS: Reference intervals were generated according to CLSI C28-A3 statistical guidelines. Caucasians, East Asians, and South Asian participants were evaluated with respect to the influence of ethnicity, and statistically significant differences were observed for 7 specific biomarkers. CONCLUSIONS: The establishment of a new comprehensive database of pediatric reference intervals is part of the Canadian Laboratory Initiative in Pediatric Reference Intervals (CALIPER). It should assist laboratorians and pediatricians in interpreting test results more accurately and thereby lead to improved diagnosis of childhood diseases and reduced patient risk. The database will also be of global benefit once reference intervals are validated in transference studies with other analytical platforms and local populations, as recommended by the CLSI.

Pediatric reference intervals for lymphocyte vitamin C (ascorbic acid).

OBJECTIVE: To establish pediatric reference intervals for lymphocyte vitamin C. DESIGN AND METHODS: This was a prospective study of 194 well children aged 0-7 years old of mixed ethnicity who had blood drawn for the purpose of this study. Blood was collected during elective surgery under general anesthesia and lymphocytes isolated and stored as frozen ascorbic acid lymphocyte lysates for later HPLC analysis by previously described methodology. Reference intervals were established according to the Clinical and Laboratory Standards Institute (CLSI) and the International Federation of Clinical Chemistry (IFCC) guidelines (C28-A3). Horn-Pesce robust method was used to estimate the 95% confidence interval and 95% reference interval. RESULTS: Reference intervals were independent of age or gender and shown to be 12.9-52.8 µg/10(8) cells (lymphocytes). CONCLUSION: We have defined pediatric reference ranges for lymphocyte vitamin C in healthy, fasted children at a relevant age group (0-7 years). The new reference interval can now be used to more reliably explore possible implications of variation of vitamin C levels on bleeding and other clinical signs.