Determination of serum ionized calcium concentration in dairy cattle after frozen anaerobic storage

The stability of serum ionized calcium concentration (ICa) from dairy cows was studied after anaerobic collection and frozen storage. Paired blood samples were obtained from five groups of cows: nonlactating, first third of lactation, midlactation, last third of lactation, and 2-year-old nonlactating heifers. Vacutainer multiple sample needles and serum separator tubes (SST) were used for venipuncture. Aspiration of serum was within 1.5 hours after collection: one sample for immediate determination (within 2 hours of collection); the other sample stored at -4 degrees C in evacuated plastic vacutainer tubes filled with serum to provide dead space of less than 75% of volume, and analyzed after 14 to 30 days in storage (half, 15, of the samples from each lactation group were analyzed after frozen anaerobic storage at 14 or 30 days, respectively). Processing samples in this manner significantly altered the values obtained for ICa, normalized calcium concentration (NCa), and pH. Analysis after frozen storage in evacuated tubes caused ICa and NCa concentrations to decrease and pH to increase (P< 0.05); total calcium levels were not significantly different from initial values. There were no significant differences among lactation groups. The difference between values obtained from these paired samples was either due to loss of CO(2) during transfer from the SST to the evacuated tube or during frozen storage. Changes in samples assayed after freezing and storage could be adjusted to original values by using the mean difference between the fresh and frozen levels as correction factors: ICa (+0.4379), NCa (+0.2797), and pH (-0.0926). It was concluded that immediate determination of serum ICa in dairy cattle is the ideal but using this methodology and performing analyses later may be acceptable if correction factors are determined.