A blood sample was obtained for laboratory analyses from all but one child. Local anaesthetical patches (EMLA R; AstraZeneca AB, Södertälje, Sweden) were used to reduce the discomfort of venipuncture. Dietary intakes were calculated from 3-day food records with Diet32 software (Aivo Oy Finland, Turku, Finland). The buy 4SC-202 nutrient contents of the foods was based
on the Finnish National Food Composition Database, Fineli, version 2001, maintained by the National Public 3-Methyladenine manufacturer Health Institute of Finland, Nutrition Unit. The total intake of vitamin D included intake from diet and from supplements. Laboratory measurements Serum 25-OHD was measured with an OCTEIA immunoenzymometric assay (IDS, Bolton, UK). The intra-assay coefficient of variation (CV) was less than 3.9% and interassay variation (4.5%). Reproducibility was ensured by adhering to the Vitamin D External Quality Assessment Scheme (DEQAS). EIA SB-715992 results were compared with HPLC results in order to determine the reliability of EIA in measuring 25-OHD2 concentration. The results were consistent (r = 0.751, p < 0.001, R 2 = 0.495); therefore, the EIA results were used throughout the study. Vitamin D status in children was defined as deficient when S-25-OHD was below 37.5 nmol/l, insufficient when it was between 37.6 and 50 nmol/l,
and sufficient when it was above 50 nmol/l, according to the published pediatric reference values [20]. In adults, a concentration of at least 80 nmol/l is considered optimal for multiple health outcomes [22]. Serum bone-specific alkaline phosphatase (S-BALP) was assayed with an OCTEIA Octase BAP immunoenzymometric assay (IDS) in order to characterize bone formation. Samples were diluted 1:5 to meet the standard curve. Intra- and interassay CVs were 6.1% and 6.7%, respectively. The bone resorption marker, serum active isoform 5b of the tartrate-resistant acid phosphatase (S-TRACP), was determined with a bone TRAP assay (SBA Sciences, Turku, Finland). Intra- and interassay
CVs were 1.2% and 3.0%, respectively. pQCT bone measurement Peripheral bone variables were determined by pQCT from the left tibia. One 2.5-mm slice (voxel size, 0.4 mm) at the 20% site of distal tibia, was measured with a XCT-2000 scanner (Stratec, click here Pforzheim, Germany) as described previously [10]. Data was analyzed using version 5.50 of the manufacturer’s software package, in which the bone contour was analyzed with a single threshold of 180 mg/cm3 for the detection of total bone mineral density (BMD), BMC, and CSA. The long-term CVs for the phantom BMD and CSA were 1.9% and 1.1%, 2.7% and 0.79%, and 0.50% and 0.78% in the total, cortical, and trabecular bone, respectively. Short-term precision (CV%) was determined with duplicate measurements of five subjects. CVs for the total bone BMD and CSA were 6.0% and 6.5%, respectively. On this basis, the calculated least significant changes for total bone BMD and CSA were 16.7% and 18.1%, respectively.