Effects of sample collection and handling on concentration of osteocalcin in equine serum

Esther Hope From the Departments of Veterinary Diagnostic Medicine (Hope, Murphy), Small Animal Clinical Sciences (Johnston, Hegstad), and Clinical and Population Sciences (Geor), College of Veterinary Medicine, University of Minnesota, St. Paul, MN 55108.

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Shirley D. Johnston From the Departments of Veterinary Diagnostic Medicine (Hope, Murphy), Small Animal Clinical Sciences (Johnston, Hegstad), and Clinical and Population Sciences (Geor), College of Veterinary Medicine, University of Minnesota, St. Paul, MN 55108.

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Rebecca L. Hegstad From the Departments of Veterinary Diagnostic Medicine (Hope, Murphy), Small Animal Clinical Sciences (Johnston, Hegstad), and Clinical and Population Sciences (Geor), College of Veterinary Medicine, University of Minnesota, St. Paul, MN 55108.

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Raymond J. Geor From the Departments of Veterinary Diagnostic Medicine (Hope, Murphy), Small Animal Clinical Sciences (Johnston, Hegstad), and Clinical and Population Sciences (Geor), College of Veterinary Medicine, University of Minnesota, St. Paul, MN 55108.

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Michael J. Murphy From the Departments of Veterinary Diagnostic Medicine (Hope, Murphy), Small Animal Clinical Sciences (Johnston, Hegstad), and Clinical and Population Sciences (Geor), College of Veterinary Medicine, University of Minnesota, St. Paul, MN 55108.

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Summary

A commercially available radioimmunoassay kit for measurement of human osteocalcin was validated for use in horses. For accurate measurement of equine serum osteocalcin, blood samples may be collected at a temperature between 20 and 25 C, then centrifuged within 90 minutes; serum may be stored at —20 C in plastic tubes for up to 26 weeks. Serum may be thawed and refrozen up to 5 times without significant change in measured equine serum osteocalcin concentration. Assay sensitivity was 0.16 ng/ml. Recovery of bovine osteocalcin standard added to equine serum was linear. Intra-assay coefficient of variation (X 100) for 2 equine serum pools was 6.9 (mean ± SD, 13.9 ± 1.0 ng/ml) and 7.5 (10.6 ± 0.8 ng/ml) %. Interassay coefficient of variation for 3 equine serum pools measured in 12 assays was 12.5 (16.1 ± 2.0 ng/ml), 12.7 (11.5 ± 1.5 ng/ml), and 24.6 (3.0 ± 0.7 ng/ml) %. Dilutional parallelism was documented by assaying pooled equine serum at 4 dilutions and correcting the mean result for dilution. Significant change was not observed in equine serum osteocalcin concentration for various time-of-day blood sample collections in horses housed under continuous lighting.

Summary

A commercially available radioimmunoassay kit for measurement of human osteocalcin was validated for use in horses. For accurate measurement of equine serum osteocalcin, blood samples may be collected at a temperature between 20 and 25 C, then centrifuged within 90 minutes; serum may be stored at —20 C in plastic tubes for up to 26 weeks. Serum may be thawed and refrozen up to 5 times without significant change in measured equine serum osteocalcin concentration. Assay sensitivity was 0.16 ng/ml. Recovery of bovine osteocalcin standard added to equine serum was linear. Intra-assay coefficient of variation (X 100) for 2 equine serum pools was 6.9 (mean ± SD, 13.9 ± 1.0 ng/ml) and 7.5 (10.6 ± 0.8 ng/ml) %. Interassay coefficient of variation for 3 equine serum pools measured in 12 assays was 12.5 (16.1 ± 2.0 ng/ml), 12.7 (11.5 ± 1.5 ng/ml), and 24.6 (3.0 ± 0.7 ng/ml) %. Dilutional parallelism was documented by assaying pooled equine serum at 4 dilutions and correcting the mean result for dilution. Significant change was not observed in equine serum osteocalcin concentration for various time-of-day blood sample collections in horses housed under continuous lighting.

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