Optimization of sample handling and processing for the carbon 13-labeled aminopyrine demethylation blood test and determination of a reference range for test results in healthy dogs.

OBJECTIVE

To determine the optimal sample handling and processing conditions for the carbon 13 ((13)C)-labeled aminopyrine demethylation blood test (ADBT; phase 1) and determine the reference range for test results (phase 2) in apparently healthy dogs.

ANIMALS

44 apparently healthy dogs (phase 1, 19 dogs; phase 2, 44 dogs).

PROCEDURES

In phase 1, a blood sample from each dog was collected before and 45 minutes after (day 0) IV administration of (13)C-labeled aminopyrine (2 mg/kg); aliquots were immediately transferred into tubes containing sodium heparin and hydrochloric acid (samples A and B), sodium heparin alone (samples C, D, and E), or sodium fluoride (sample F). Hydrochloric acid was added to samples C through F at days 7, 14, 21, and 21, respectively. The baseline and 45-minute samples' absolute (13)C:(12)C ratios were determined via fractional mass spectrometry on day 0 (control sample A) or 21 (samples B through F) and used to calculate the percentage dose of (13)C recovered in CO(2) extracted from samples (PCD). In phase 2, blood samples from each dog were collected into tubes containing sodium fluoride and processed within 3 weeks.

RESULTS

Compared with the control sample value, PCDs for samples C through E differed significantly, whereas PCD in sample F did not. The (13)C-ADBT-derived PCD reference range (central 95th percentile) for apparently healthy dogs was 0.08% to 0.2%.

CONCLUSIONS AND CLINICAL RELEVANCE

Glycolytic CO(2) production in canine blood samples collected during (13)C-ADBTs was sufficiently inhibited by sodium fluoride to allow delayed sample analysis and avoid transportation of hydrochloric acid-treated samples.