OBJECTIVE To investigate effects of storage duration and temperature on biochemical analytes in plasma from red-eared sliders (Trachemys scripta elegans).
ANIMALS 8 red-eared sliders.
PROCEDURES Blood samples were collected. Plasma was harvested and analyzed at room temperature (approx 23°C; time = 1 hour) and then fractioned into 0.1-mL aliquots that were stored at room temperature or were refrigerated (4°C) or frozen (−20°C). Biochemical analysis of stored samples was performed at 4 (room temperature), 8 (4°C), 24 (4°C), 48 (4° and −20°C), and 72 (−20°C) hours and at 7 days (−20°C). For each time point for each storage temperature, bias was calculated by subtracting values from the value obtained at 1 hour. Bias was modeled by use of a linear mixed model.
RESULTS Storage temperature had a significant effect on several plasma biochemical analytes. In general, aspartate aminotransferase activity and uric acid, total protein, and potassium concentrations increased after storage at 4° and −20°C. Differences in values after storage were mostly within the acceptable range for allowable total error, except for calcium and potassium concentrations for samples stored at −20°C. Both storage temperatures increased variability of measurement results. Results for samples stored at room temperature for 4 hours did not differ significantly from values at 1 hour. Results differed significantly between refrigerated and frozen samples stored for 48 hours.
CONCLUSIONS AND CLINICAL RELEVANCE Short-term storage conditions influenced results for some biochemical analytes. These effects should be considered when performing biochemical analyses of plasma samples obtained from red-eared sliders.
OBJECTIVE To evaluate effects of blood contamination on dipstick results, specific gravity (SG), and urine protein-to-urine creatinine ratio (UPCR) for urine samples from dogs and cats.
SAMPLE Urine samples collected from 279 dogs and 120 cats.
PROCEDURES Urine pools were made for each species (dogs [n = 60] and cats ). Blood was added to an aliquot of a pool, and serial dilutions were prepared with the remaining urine. Color and dipstick variables were recorded, and SG and UPCR were measured. For cats, 1 set of pools was used; for dogs, 2 sets were used. Comparisons were made between undiluted urine and spiked urine samples for individual colors. Repeated-measures ANOVA on ranks was used to compare dipstick scores and UPCR results; χ2 tests were used to compare proteinuria categorizations (nonproteinuric, borderline, or proteinuric).
RESULTS Any blood in the urine resulted in significantly increased dipstick scores for blood. In both species, scores for bilirubin and ketones, pH, and SG were affected by visible blood contamination. No significant difference for the dipstick protein reagent results was evident until a sample was visibly hematuric. The UPCR was significantly increased in dark yellow samples of both species. Proteinuria categorizations differed significantly between undiluted urine and urine of all colors, except light yellow.
CONCLUSIONS AND CLINICAL RELEVANCE Any degree of blood contamination affected results of dipstick analysis. Effects depended on urine color and the variable measured. Microscopic blood contamination may affect the UPCR; thus, blood contamination may be a differential diagnosis for proteinuria in yellow urine samples.
OBJECTIVE To assess the effect of decreased platelet and WBC counts on platelet aggregation as measured by a multiple-electrode impedance aggregometer in dogs.
ANIMALS 24 healthy dogs.
PROCEDURES From each dog, 9 mL of blood was collected into a 10-mL syringe that contained 1 mL of 4% sodium citrate solution to yield a 10-mL sample with a 1:9 citrate-to-blood ratio. Each sample was then divided into unmanipulated and manipulated aliquots with progressively depleted buffy-coat fractions such that 2 to 3 blood samples were evaluated per dog. The Hct for manipulated aliquots was adjusted with autologous plasma so that it was within 2% of the Hct for the unmanipulated aliquot for each dog. All samples were analyzed in duplicate with a multiple-electrode impedance aggregometer following the addition of ADP as a platelet agonist. The respective effects of platelet count, plateletcrit, Hct, and WBC count on platelet aggregation area under the curve (AUC), aggregation, and velocity were analyzed with linear mixed models.
RESULTS WBC count was positively associated with platelet AUC, aggregation, and velocity; blood samples with leukopenia had a lower AUC, aggregation, and velocity than samples with WBC counts within the reference range. Platelet count, plateletcrit, and Hct did not have an independent effect on AUC, aggregation, or velocity.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that WBC count was positively associated with platelet aggregation when ADP was used to activate canine blood samples for impedance aggregometry. That finding may be clinically relevant and needs to be confirmed by in vivo studies.
OBJECTIVE To provide contemporary preliminary guidelines for the morphological evaluation of bone marrow in conjunction with CBC results for healthy juvenile (3- to 6-month-old) female New Zealand White rabbits (Oryctolagus cuniculus).
ANIMALS 22 female New Zealand White rabbits.
PROCEDURES Each rabbit was sedated, and a blood sample (3 mL) was collected from an ear artery for a CBC, after which the rabbit was euthanized. Within 5 minutes after euthanasia, bone marrow samples were obtained from the femur for cytologic and histologic evaluation. Bone marrow specimens for cytologic evaluation were stained with modified Wright stain, and those for histologic evaluation were stained with either H&E or Prussian blue stain.
RESULTS The CBC results were within published reference ranges for all rabbits except 4, each of which had mild leukopenia. Cytologic assessment of bone marrow revealed a median myeloid-to-erythroid ratio of 0.7 and 2.8 megakaryocytes/low-power field (magnification, 100X), and the median percentages of lymphocytes, plasma cells, and macrophages were 11.5%, 0.1%, and 0%, respectively. The myeloid-to-erythroid ratio was not significantly correlated with any CBC variable. On histologic evaluation of bone marrow, the cellularity ranged from 30% to 50%, there were 2.1 to 7.7 megakaryocytes/hpf (magnification, 400X), and no iron stores were visible in H&E or Prussian blue–stained specimens.
CONCLUSIONS AND CLINICAL RELEVANCE Results of the present study provided contemporary preliminary guidelines for the evaluation of bone marrow in healthy laboratory rabbits.
OBJECTIVE To purify and characterize equine vitamin D-binding protein (VDBP) from equine serum and to evaluate plasma concentrations of VDBP in healthy horses and horses with gastrointestinal injury or disease.
ANIMALS 13 healthy laboratory animals (8 mice and 5 rabbits), 61 healthy horses, 12 horses with experimentally induced intestinal ischemia and reperfusion (IR), and 59 horses with acute gastrointestinal diseases.
PROCEDURES VDBP was purified from serum of 2 healthy horses, and recombinant equine VDBP was obtained through a commercial service. Equine VDBP was characterized by mass spectrometry. Monoclonal and polyclonal antibodies were raised against equine VDBP, and a rocket immunoelectrophoresis assay for equine VDBP was established. Plasma samples from 61 healthy horses were used to establish working VDBP reference values for study purposes. Plasma VDBP concentrations were assessed at predetermined time points in horses with IR and in horses with naturally occurring gastrointestinal diseases.
RESULTS The working reference range for plasma VDBP concentration in healthy horses was 531 to 1,382 mg/L. Plasma VDBP concentrations were significantly decreased after 1 hour of ischemia in horses with IR, compared with values prior to induction of ischemia, and were significantly lower in horses with naturally occurring gastrointestinal diseases with a colic duration of < 12 hours than in healthy horses.
CONCLUSIONS AND CLINICAL RELEVANCE Plasma VDBP concentrations were significantly decreased in horses with acute gastrointestinal injury or disease. Further studies and the development of a clinically relevant assay are needed to establish the reliability of VDBP as a diagnostic and prognostic marker in horses.
OBJECTIVE To compare platelet function and viscoelastic test results between healthy dogs and dogs with chronic kidney disease (CKD) to assess whether dogs with CKD have platelet dysfunction and altered blood coagulation.
ANIMALS 10 healthy control dogs and 11 dogs with naturally occurring CKD.
PROCEDURES Blood and urine were collected once from each dog for a CBC, serum biochemical analysis, urinalysis, and determination of the urine protein-to-creatinine ratio, prothrombin time, activated partial thromboplastin time, plasma fibrinogen concentration, and antithrombin activity. Closure time was determined by use of a platelet function analyzer and a collagen-ADP platelet agonist. Thromboelastography (TEG) variables (reaction time, clotting time, α angle, maximum amplitude, and global clot strength [G value]) were determined by use of recalcified nonactivated TEG. Platelet expression of glycoprotein Ib (GPIb; receptor for von Willebrand factor), integrin αIIbβ3 (αIIbβ3; receptor for fibrinogen), and P-selectin (marker for platelet activation) was assessed by flow cytometry.
RESULTS Compared with healthy control dogs, the median closure time was prolonged, the median maximum amplitude and G value were increased, and the median clotting time was decreased for dogs with CKD. Platelet expression of both αIIbβ3 and P-selectin was also significantly increased for dogs with CKD, compared with that for control dogs. Platelet expression of GPIb, αIIbβ3, and P-selectin was not correlated with closure time or any TEG variable.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that dogs with CKD frequently had evidence of platelet dysfunction and hypercoagulability that were not totally attributable to alterations in platelet surface expression of GPIb, αIIbβ3, and P-selectin.
OBJECTIVE To establish reference intervals for serum reactive oxygen metabolites (ROMs), biological antioxidant potential (BAP), and oxidative stress index (OSi) in adult rams by use of controlled preanalytic and analytic procedures.
ANIMALS 123 healthy 1- to 4-year-old rams of 2 Greek breeds (Chios [n = 62] and Florina ).
PROCEDURES 4 hours after rams were fed, a blood sample was obtained from each ram, and serum was harvested. Concentrations of ROMs and BAP were measured colorimetrically on a spectrophotometric analyzer. The OSi was calculated as ROMs concentration divided by BAP concentration. Combined and breed-specific reference intervals were calculated by use of nonparametric and robust methods, respectively. Reference intervals were defined as the 2.5th to 97.5th percentiles.
RESULTS Reference intervals for ROMs, BAP, and OSi for all rams combined were 65 to 109 Carratelli units, 2,364 to 4,491 μmol/L, and 18.2 to 43.0 Carratelli units/(mmol/L), respectively. Reference intervals of Chios rams for ROMs, BAP, and OSi were 56 to 113 Carratelli units, 2,234 to 4,290 μmol/L, and 12.9 to 38.4 Carratelli units/(mmol/L), respectively. Reference intervals of Florina rams for ROMs, BAP, and OSi were 68 to 111 Carratelli units, 2,337 to 4,363 μmol/L, and 14.1 to 38.1 Carratelli units/(mmol/L), respectively.
CONCLUSIONS AND CLINICAL RELEVANCE Reference intervals calculated in this study can be used as a guide for the interpretation of ROMs, BAP, and OSi results in rams and, under appropriate conditions, can be adopted for use by veterinary laboratories.
OBJECTIVE To determine the predominant thromboxane (TX) metabolite in urine of healthy cats, evaluate whether the method of sample collection would impact concentration of that metabolite, and propose a reference interval for that metabolite in urine of healthy cats.
ANIMALS 17 cats (11 purpose-bred domestic shorthair cats, 5 client-owned domestic shorthair cats, and 1 client-owned Persian cat).
PROCEDURES All cats were deemed healthy on the basis of results for physical examination, a CBC, serum biochemical analysis, urinalysis, and measurement of prothrombin time and activated partial thromboplastin time. Voided and cystocentesis urine samples (or both) were collected. Aliquots of urine were stored at −80°C until analysis. Concentrations of TXB2, 11-dehydroTXB2, and 2,3 dinorTXB2 were measured with commercially available ELISA kits. Urinary creatinine concentration was also measured.
RESULTS 11-dehydroTXB2 was the most abundant compound, representing (mean ± SD) 59 ± 18% of the total amount of TX detected. In all samples, the concentration of 11-dehydroTXB2 was greater than that of 2,3 dinorTXB2 (mean, 4.2 ± 2.7-fold as high). Mean concentration of 11-dehydroTXB2 for the 17 cats was 0.57 ± 0.47 ng/mg of creatinine. A reference interval (based on the 5% to 95% confidence interval) of 0.10 to 2.1 ng of 11-dehydroTXB2/mg of creatinine was proposed for healthy cats.
CONCLUSIONS AND CLINICAL RELEVANCE In this study, 11-dehydroTXB2 was the major TX metabolite in feline urine. Measurement of this metabolite may represent a noninvasive, convenient method for monitoring in vivo platelet activation in cats at risk for thromboembolism.
OBJECTIVE To evaluate use of serum amyloid A (SAA) and haptoglobin concentrations as prognostic indicators for horses with inflammatory disease in regard to euthanasia, complications, and hospitalization duration and cost.
ANIMALS 20 clinically normal horses and 53 horses with inflammatory disease.
PROCEDURES Total WBC count, neutrophil count, and fibrinogen, SAA, and haptoglobin concentrations were determined for clinically normal horses and horses with suspected inflammatory disease. Clinicopathologic values at admission were compared to test the use of SAA and haptoglobin concentrations in predicting euthanasia, complications, and hospitalization duration and cost. Haptoglobin and SAA concentrations of 22 horses were monitored during hospitalization to test the use of serial measurements in predicting survival and complications.
RESULTS Neutrophil count and SAA and haptoglobin concentrations were significantly different at admission for horses with inflammatory disease, compared with those for clinically normal horses. Horses with colitis and peritonitis had significantly higher SAA and haptoglobin concentrations than clinically normal horses. A moderate positive correlation (r = 0.355) between hospitalization duration and haptoglobin concentration was identified. Horses with an increase in SAA concentration between 24 and 72 hours after admission, compared with admission SAA concentration, were significantly more likely (OR, 7.0; 95% confidence interval, 1.1 to 45.9) to be euthanized or develop complications.
CONCLUSIONS AND CLINICAL RELEVANCE Concentrations of SAA and haptoglobin at admission were not significantly correlated with outcome in horses with inflammatory conditions. Acute-phase proteins likely have more utility in serial analysis rather than testing at a single time point for horses with inflammatory conditions.
OBJECTIVE To compare blood glucose concentrations of black-tailed prairie dogs (Cynomys ludovicianus) measured by use of a variety of portable analyzers with results from a laboratory biochemistry analyzer.
SAMPLE Venous blood samples (3 mL) obtained from each of 16 healthy black-tailed prairie dogs.
PROCEDURES A portion of each blood sample was used to measure glucose concentrations by use of an amperometric human point-of-care glucometer and a colorimetric species-specific portable blood glucose meter designed for veterinary use with both canine (code 5) and feline (code 7) settings. The remainder of each blood sample was placed into 2 tubes (one contained lithium heparin and the other contained no anticoagulant). A portable veterinary chemistry analyzer (PVCA) and a handheld analyzer were used to measure glucose concentration in heparinized blood. Serum glucose concentration was measured in the remaining portion by use of a biochemistry analyzer. A general linear mixed models approach was used to compare glucose concentrations and measurement bias obtained with the various measurement methods.
RESULTS Measurement bias and differences in mean glucose concentrations were apparent with all measurement methods. In particular, the veterinary glucometer, whether used on the canine or feline setting, overestimated mean glucose concentrations, whereas the human glucometer, PVCA, and handheld analyzer underestimated mean glucose concentrations relative to the concentration obtained with the biochemistry analyzer.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that none of the measurement methods provided consistently accurate blood glucose concentrations of black-tailed prairie dogs, compared with values determined with a biochemistry analyzer.