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

The research article discusses the validation of a commercially available test kit, originally developed for humans, for measuring osteocalcin levels in horse serum. It suggests that accurate measures can be achieved if blood samples are collected at specific conditions, handled and stored properly.

Validation of Radioimmunoassay Kit

• The study focused on validating a commercially available radioimmunoassay kit meant for measuring levels of osteocalcin in humans for its use in horses.
• Osteocalcin is a protein produced by bone-building cells (osteoblasts) and its levels can be an indicator of bone health and metabolic disorders.
• Radioimmunoassay (RIA) is a sensitive and specific method for measuring antigens (like osteocalcin). The process involves competition between radiolabeled antigen and unlabeled antigen for a fixed number of antibody binding sites.

Sample Collection and Handling

• In order to accurately measure equine serum osteocalcin, blood samples need to be collected at a temperature between 20 and 25 degrees Celsius, then centrifuged within 90 minutes.
• Serum samples can then be stored at -20 degrees Celsius in plastic tubes for up to 26 weeks.
• Additionally, serum samples can be thawed and refrozen up to 5 times without any significant change in the measured equine serum osteocalcin concentration.

Experimental Results

• The assay sensitivity was found to be 0.16 ng/ml, meaning it could detect osteocalcin concentrations as low as this.
• The recovery of bovine osteocalcin standard when added to equine serum was linear, implying that the assay provided consistent results across a range of osteocalcin concentrations.
• Conducting a series of intra-assay and interassay tests further validated the method’s accuracy and precision.
• Dilutional parallelism was documented, which suggested that the signals produced by different dilutions of a sample were proportional, indicating consistent accuracy of the assay over a range of concentrations.
• No significant change was observed in equine serum osteocalcin concentration when blood samples were collected at different times of day in horses under continuous lighting, indicating that the test results were not influenced by the time of day the sample was taken.