Quality of hematology and clinical chemistry results in laboratory and zoo nonhuman primates: Effects of the preanalytical phase. A review.

Most errors in clinical pathology originate in the preanalytical phase, which includes all steps from the preparation of animals and equipment to the collection of the specimen and its management until analyzed. Blood is the most common specimen collected in nonhuman primates. Other specimens collected include urine, saliva, feces, and hair. The primary concern is the variability of blood hematology and biochemistry results due to sampling conditions with the effects of capture, restraint, and/or anesthesia. Housing and diet have fewer effects, with the exception of food restriction to reduce obesity. There has been less investigation regarding the impact of sampling conditions of nonblood specimens.

Improving the Quality of EDTA-treated Blood Specimens from Mice.

Nonterminal blood sampling in laboratory mice is a very common procedure. With the goal of improving animal welfare, different sampling sites and methods have been compared but have not achieved a consensus. Moreover, most of these studies overlooked the quality of blood specimens collected. The main preanalytical concern with EDTA-treated blood specimens for hematology analyses is platelet aggregation, which is known to cause analytical errors. Our objective was to find a nonterminal blood sampling method with minimal adverse effects on mice and few or no platelet aggregates. We tested and compared 2 collection sites, 4 sampling methods, and 3 antithrombotic drugs in 80 C57BL6/j male and female mice by evaluating platelet aggregates on blood smears and platelet, WBC, and RBC counts. In addition, the blood collection process was carefully evaluated, and adverse effects were recorded. Platelet aggregation was lower in specimens collected from the jugular vein than from the facial vein, with no effect of the sampling device or the presence of an antithrombotic additive. Highly aggregated specimens were significantly associated with lower platelet counts, whereas aggregation had no effect on WBC or RBC counts. Adverse events during sampling were significantly associated with more numerous platelet aggregates. The jugular vein is thus a satisfactory sampling site in mice in terms of both animal welfare and low platelet aggregation. Using antithrombotic agents appears to be unnecessary, whereas improving sampling conditions remains a key requirement to ensure the quality of EDTA-treated blood specimens from mice.

Validation of the Sysmex XN-V hematology analyzer for canine specimens.

BACKGROUND: The Sysmex XN-V is derived from the new Sysmex XN series of human hematology analyzers. The main changes from the previously validated XT-2000iV analyzer include an optic-fluorescent analysis for platelets and nucleated RBC count. OBJECTIVE: We aimed to validate the Sysmex XN-V for canine blood according to American College for Veterinary Clinical Pathology and International Council for Standardization in Hematology recommendations. MATERIALS AND METHODS: Canine EDTA blood specimens and quality control material were analyzed on the Sysmex XN-V to evaluate imprecision, bias, linearity, a comparison with the XT-2000iV analyzer, interference effects, carry-over, and stability. We also verified previously established Sysmex XT-2000iV reference intervals (RIs). RESULTS: Imprecision and bias were low (<5%) for most variables. Observed total error was lower than allowable total error for most measured variables except lymphocytes and monocytes. Visually determined linearity was excellent for all variables, except for lymphocytes. The correlation between the XN-V and XT-2000iV analyzers was high (>0.93) for all variables except MCHC and reticulocyte indices. Correlations between the Sysmex XN-V and manual differential counts were good for neutrophils and eosinophils, acceptable for lymphocytes, and fair for monocytes. Hemolysis, lipemia, and to a lesser extent icterus, had significant effects on measured hemoglobin concentration and associated variables. Carry-over was not visually observed for any variable. Changes in the Sysmex XN-V measurements after storage at 4℃ and 24℃ were similar to those described for the Sysmex XT-2000iV analyzer. The previously established Sysmex XT-2000iV RIs can be used to interpret results from the Sysmex XN-V analyzer for most variables except red blood cell distribution width and mean platelet volume. CONCLUSIONS: The performance of the Sysmex XN-V analyzer was excellent and compared favorably with the Sysmex XT-2000iV analyzer.

Reference Values for Hematology, Plasma Biochemistry, Bone Marrow Cytology and Bone Histology of NOD.Cg-Prkdcscid Il2rgtm1Wjl/ SzJ Immunodeficient Mice.

Highly immunodeficient NSG mice (NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ) are commonly used as a models in preclinical studies for patient-derived engraftment. However, despite the frequency of their use, reference values for their clinical pathology markers have not been determined. In accordance with the American Society of Veterinary Clinical Pathology (ASVCP) recommendations, we established de novo reference values for hematologic and biochemical variables and evaluated bone marrow cytology and histology in forty 9-wk-old male and female NSG mice. Hematologic analyses were performed using 2 separate analyzers (IDEXX ProCyte Dx, Sysmex XT-2000iV) and biochemical values were measured using a Scil VetScan2. The primary hematologic characteristic seen in NSG mice was a very low white blood cell (WBC) count (below 1.6 109/L). Lymphocyte and monocyte counts were respectively over- and under-estimated by the analyzers, as compared with manual counts, likely due to misidentification of the very low concentrations of these cell types by the analyzers. This analytical bias highlights the need for confirmatory microscopic observation of blood smears from these mice for WBC differential identification. Results for all other hematology and biochemistry variables were similar to those previously reported in inbred mice, except for MPV and an unexpectedly high glucose concentration (11.5 to 19.0 mmol/L), potentially due to the nonfasting status of the animals. The differential bone marrow cell count and Myeloid:Erythroid ratio (median 1.76) were also established. Megakaryocyte and adipocyte count differed significantly between the femoral diaphysis and metaphysis and between genders. These results provide a reliable resource of baseline data for hematologic variables for researchers monitoring graft rejection studies in NSG mice.

The effects of storage at 4°C and 20°C on the hemograms of C57BL/6 mice and Wistar rats using the IDEXX ProCyte Dx and blood smear evaluations.

BACKGROUND: Delayed blood analysis might be unavoidable in laboratory practice, but little is known about rodent blood stability, especially cell morphology and scattergram results. OBJECTIVES: This study aimed to evaluate the stability of rodent blood cell counts and morphologies at different temperatures using the ProCyte Dx analyzer and performing manual observations. METHODS: Ten Wistar rats and 10 C57bl/6 mice were sampled on EDTA tubes and aliquoted for storage (4°C, 20°C). Hematologic analyses were performed immediately and at T6h, T24h, T48h (rats and mice), and T72h (rats only) after storage. RESULTS: In rats, at any temperature, red blood cell counts, hemoglobin concentrations (HGB), mean corpuscular hemoglobin (MCH) levels, and reticulocyte, white blood cell (WBC), eosinophil, and impedance platelet counts remained stable over time. The main changes were observed at 20°C for hematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), and WBC differential counts. Optical platelet counts (PLT-O) and platelet variables underwent changes at both temperatures from T24h. In mice, red blood cell counts by impedance (RBC-I), MCH, and WBC, lymphocyte, eosinophil, and platelet counts, and plateletcrit (PCT) were stable over time and at all temperatures. As in rats, the most significant changes were observed at 20°C and concerned the optical RBC (RBC-O) counts, HCTs, MCVs, MCHCs, and reticulocyte, neutrophil, and monocyte counts. For both species, blood cell morphologies were altered from T24h at all temperatures, and platelet clumps were more numerous at 4°C. CONCLUSIONS: When rodent blood analyses need to be delayed, storage at 4°C is preferred and should not exceed 24 hours. PLT counts should be interpreted cautiously in refrigerated specimens with mandatory blood smear evaluations when abnormal scattergrams are observed.

Urinalysis and determination of the urine protein-to-creatinine ratio reference interval in healthy cows.

BACKGROUND: There are no reference intervals for urinalysis in cattle. HYPOTHESIS/OBJECTIVES: Characterize the urine of healthy cows, establish urine protein-to-creatinine ratio (UPC) reference intervals, and test possible differences among dairy and beef cattle, age groups, or stage of lactation. ANIMALS: Seventy-seven dairy and 74 beef 2.5 to 17 year-old cows of different breeds housed mainly in free stall. METHODS: In this prospective study, urine specimens were collected by catheterization. Complete urinalysis was performed within 1 hour including specific gravity, dipstick evaluation, visual urine pH evaluation with 0.3 pH unit graded strips, and microscopic evaluation of the sediment. Urinary protein and creatinine concentrations and protein electrophoresis were determined on frozen aliquots. RESULTS: Overall reference intervals were 1.020 to 1.045 for USG, 7.0 to 8.7 for pH, and 0.04 to 0.25 for UPC; because of differences in creatinine concentration, UPC was lower in beef (0.04-0.14) than in dairy (0.05-0.25) cows and in the latter in dry than lactating cows. With dipstick evaluation, most analytes were absent except for blood, ketone, and protein in 24.7, 16.0, and 64.7% of cases, respectively. Microscopic evaluation revealed less than 3 red blood cells, leukocytes, and epithelial cells in 84, 99.3, and 100% cows, respectively. No band was observed at electrophoresis, except in 1 case at MW ~66 000. CONCLUSIONS AND CLINICAL IMPORTANCE: Creatininuria is higher in beef than dairy cows and proteinuria is likely more efficiently characterized by protein concentration than by UPC.

Hematology reference intervals for adult cows in France using the Sysmex XT-2000iV analyzer.

In order to develop bovine hematology reference intervals (RIs) in accordance with new international recommendations, we analyzed 156 blood specimens of healthy adult dairy and beef cows from 32 farms with a Sysmex XT-2000iV analyzer, and by manual scoring of platelet clumps and white blood cell (WBC) differential. We established RIs by the nonparametric method, and examined effects of age, production type (beef vs. dairy), and stage of lactation. RIs could not be determined for platelet count and indices because clumps were observed in 80% of specimens. Optical and impedance red blood cell (RBC) counts were similar, although statistically different. RIs for analyzer and manual WBC differentials were not different except for lymphocyte concentration, the subpopulations of which were counted manually. Hematocrit was higher in beef than dairy cattle, and hemoglobin was lower in early lactation. Increases in RBC count, mean corpuscular volume, mean corpuscular hemoglobin, and RBC distribution width were noted with increasing age, along with decreases in WBC count, neutrophils, and lymphocytes. Most RIs in our study, with the exception of neutrophils, were similar to those previously reported using a flow cytometry analyzer.

Effects of storage conditions on results for quantitative and qualitative evaluation of proteins in canine urine.

OBJECTIVE To investigate effects of storage conditions on the canine urine protein-to-creatinine ratio (UPC) and on SDS-agarose gel electrophoresis (AGE) of urinary proteins. SAMPLE Urine specimens from 20 proteinuric (UPC > 0.5) and 20 nonproteinuric (UPC ≤ 0.2) dogs. PROCEDURES UPC and SDS-AGE were performed on urine specimens stored at room temperature (20°C) and 4°C for up to 5 days and at -20° and -80°C for up to 360 days; some specimens were subjected to 3 freeze-thaw cycles. Results were compared with those obtained for fresh urine specimens. RESULTS UPC was not affected by storage at room temperature or by freezing. A decrease in UPC was observed for specimens from nonproteinuric dogs after 5 days at 4°C (10%) and from both groups after 90 days at -20° and -80°C (≤ 20% and ≤ 15%, respectively). The SDS-AGE profiles revealed no visual changes regardless of duration of storage for specimens stored at room temperature, 4°C, and -80°C, except for 1 profile after 360 days at -80°C. Repeated freeze-thaw cycles did not affect SDS-AGE profiles. Appearance or strengthening of high-molecular-weight bands that could alter interpretation was evident in SDS-AGE profiles after storage at -20°C for ≥ 15 days (31/40 dogs). CONCLUSIONS AND CLINICAL RELEVANCE Storage of urine at -20° or -80°C for up to 1 year influenced the UPC without affecting clinical interpretation. Storage of urine specimens at -20°C impaired visual analysis of SDS-AGE. When SDS-AGE cannot be performed on fresh or recently refrigerated urine specimens, storage at -80°C is recommended.

Evaluation of CTAD (citrate-theophylline-adenosine-dipyridamole) as a universal anticoagulant in dogs.

CTAD (citrate-theophylline-adenosine-dipyridamole) has been shown to be an almost universal anticoagulant in human and feline medicine, allowing most hematology, coagulation, and biochemical analyses. Forty canine blood specimens were collected in CTAD, EDTA, heparin, and citrate for hematology, biochemistry, and coagulation analyses. CTAD partially limited platelet aggregation observed in EDTA blood smears. CTAD specimens gave similar and well-correlated results for most variables of a complete blood cell count, except for mean corpuscular volume, which was moderately higher, and mean corpuscular hemoglobin concentration, which was moderately lower in CTAD than in EDTA; reticulocyte and platelet indexes were poorly correlated. CTAD plasma gave similar results to citrate for fibrinogen, antithrombin, and D-dimers, and relatively similar results for prothrombin time, but activated partial thromboplastin time was poorly correlated. Triglycerides, cholesterol, glucose, total proteins, phosphate, iron, alanine aminotransferase, γ-glutamyl transferase, and lipase were similar and well correlated in CTAD and heparin plasmas. Urea, creatinine, albumin, alkaline phosphatase, amylase, and aspartate aminotransferase showed moderate-to-marked bias, but these variables could be measured in CTAD plasma if new reference intervals were determined. Creatine kinase activity, potassium, chloride, and total carbon dioxide measurements are not recommended in CTAD plasma. CTAD is a prospective candidate as an almost universal anticoagulant for routine hematology, some plasma coagulation, and many biochemistry variables in dogs. Definitive recommendations will require study of abnormal canine blood specimens.

Hematologic and Biochemical Biologic Variation in Laboratory Cats.

The biologic variation associated with a clinical pathology result is important to consider before reference intervals (RI) are used. Most available RI are population-based RI, in which the analytical variability, interindividual variability, and intraindividual variability are confounded. In addition, when the intraindividual variability is considerably less than the interindividual variability, a population-based RI is insufficiently sensitive to detect changes in a subject over time. Here we determined the biologic variation and reference change value (RCV) of hematologic and biochemical variables in laboratory cats. Blood specimens from 14 (7 females and 7 males) overnight-fasted laboratory cats sampled 7 times (days 1, 2, 7, 14, 31, 42, and 100) were analyzed regarding hematology and biochemistry variables. For each variable, analytical, intraindividual, and interindividual coefficients of variation were estimated prior to calculation of the index of individuality and the RCV. RBC variables (count, Hgb, Hct, MCV, MCH, MCHC, and RBC distribution width) and 5 biochemical analytes (cholesterol, creatinine, triglycerides, ALP, and calcium) exhibited marked individuality, therefore indicating that subject-based reference intervals or RCV would be preferable when monitoring these variables in laboratory cats. Population-based RI were shown to be adequate for glucose and sodium, and both types of population and individual RI were similarly efficient for albumin, total protein, urea, ALT, AST, creatine kinase, chloride, carbon dioxide, iron, magnesium, inorganic phosphate, and potassium and reticulocyte, WBC, neutrophil, lymphocyte, monocyte, eosinophil, and platelet counts. The RCV determined in the present study provide a valuable tool for monitoring hematologic and biochemical variables in healthy laboratory cats.

Characterization of Proteinuria in Dogue de Bordeaux Dogs, a Breed Predisposed to a Familial Glomerulonephropathy: A Retrospective Study.

Dogue de Bordeaux dog has been reported to be predisposed to a familial glomerulonephropathy that displays some morphological modifications reported in focal and segmental glomerulosclerosis. Prevalence of quantitatively abnormal renal proteinuria was recently reported to be 33% in this breed. The nature of the proteinuria was assessed by sodium dodecyl sulfate-agarose gel electrophoresis and determinations of urinary markers (urinary retinol-binding protein, urinary N-acetyl-ß-glucosaminidase, urinary albumin and urinary immunoglobulin G) on stored specimens. Diagnostic performances of sodium dodecyl sulfate-agarose gel electrophoresis to identify dogs with elevated urinary biomarkers were assessed. Samples from 102 adult Dogue de Bordeaux dogs (47 non-proteinuric [urine protein-to-creatinine ratio ≤ 0.2], 20 borderline-proteinuric [0.2< urine protein-to-creatinine ratio ≤ 0.5] and 35 proteinuric dogs [urine protein-to-creatinine ratio >0.5]) were used, of which 2 were suffering from familial glomerulonephropathy. The electrophoretic protein patterns, for all but one proteinuric dog, were indicative of a glomerular origin and, in all dogs, the urinary albumin concentration related to creatinine concentration and the urinary immunoglobulin G concentration related to creatinine concentration were above the upper limit of the reference interval established for the breed. Sensitivity and specificity of sodium dodecyl sulfate-agarose gel electrophoresis identifying dogs with elevated urinary albumin concentration were 94% and 92%, respectively, while diagnostic performance of sodium dodecyl sulfate-agarose gel electrophoresis in detecting dogs with elevated urinary immunoglobulin G concentration yielded sensitivity and specificity of 90% and 74%, respectively. These results suggest that all proteinuric and some borderline-proteinuric Dogue de Bordeaux dogs likely have underlying glomerular lesions and that sodium dodecyl sulfate-agarose gel electrophoresis and urinary markers might be useful to screen dogs with borderline-proteinuria. Additional investigations are warranted to assess if these findings are related to the familial glomerulonephropathy.

The preanalytic phase in veterinary clinical pathology.

This article presents the general causes of preanalytic variability with a few examples showing specialists and practitioners that special and improved care should be given to this too often neglected phase. The preanalytic phase of clinical pathology includes all the steps from specimen collection to analysis. It is the phase where most laboratory errors occur in human, and probably also in veterinary clinical pathology. Numerous causes may affect the validity of the results, including technical factors, such as the choice of anticoagulant, the blood vessel sampled, and the duration and conditions of specimen handling. While the latter factors can be defined, influence of biologic and physiologic factors such as feeding and fasting, stress, and biologic and endocrine rhythms can often not be controlled. Nevertheless, as many factors as possible should at least be documented. The importance of the preanalytic phase is often not given the necessary attention, although the validity of the results and consequent clinical decision making and medical management of animal patients would likely be improved if the quality of specimens submitted to the laboratory was optimized.

Changes in hematology measurements in healthy and diseased dog blood stored at room temperature for 24 and 48 hours using the XT-2000iV analyzer.

BACKGROUND: Changes in canine hematology measurements may occur when analyses are delayed due to shipment of specimens to a laboratory. OBJECTIVE: The purpose of this study was to report changes in hematologic variables in healthy and diseased canine blood measured with a Sysmex XT-2000iV during storage at room temperature for 24 and 48 hours. METHODS: EDTA-K3 blood samples from 42 healthy and diseased dogs were measured on a Sysmex XT-2000iV analyzer within one hour of sampling, and after storage for 24 and 48 hours at room temperature in the dark. RESULTS: Storage caused little or no change in RBC count, HGB concentration and MCH, while there was a moderate increase in HCT, MCV and reticulocytes count, and a moderate decrease in MCHC. Decreased platelet counts by impedance (PLT-I) and optical (PLT-O) measurements were associated with increased mean platelet volume (MPV), platelet-large cell ratio (P-LCR) and platelet distribution width (PDW), including a right shift in the platelet histogram and a dispersion of the platelet dot plot on the scattergram. The total and differential WBC count remained stable except for decreased monocyte counts. In the scatterplots, monocytes shifted into the lymphocyte population after 24 hours, and neutrophil population shifted to the right appearing in the eosinophil gate at 48 hours of storage. The disease status had only a small effect on storage-induced changes, and observed changes had no consequences for clinical decisions. CONCLUSIONS: Blood storage at room temperature was accompanied by moderate variations in some hematologic variables, awareness of which helps in avoiding misinterpretations.

Feline reference intervals for the Sysmex XT-2000iV and the ProCyte DX haematology analysers in EDTA and CTAD blood specimens.

Laser-based haematology analysers are routinely used in veterinary clinical pathology laboratories, and are available to practitioners. However, feline haematological reference intervals (RIs) determined according to international recommendations are, to our knowledge, not available. Furthermore, platelet count RI is difficult to establish in cats because of the frequent occurrence of platelet aggregation in blood specimens. The purpose of this study was to establish feline haematological RIs with the Sysmex XT-2000iV and ProCyte DX analysers, in ethylenediamine tetra-acetic acid (EDTA) and in citrate, theophylline, adenosine and dipyridamole (CTAD), which is a combination of anticoagulants limiting platelet aggregation. Blood specimens from 120 healthy cats were analysed in duplicate, and the degree of platelet aggregation was assessed on blood smears. After exclusion of inadequate specimens, 81 sets of results (from 44 males and 37 females, aged from 6 to 116 months) were available for the determination of RIs by the non-parametric method. The effects of the anticoagulant, analyser and aggregation score were assessed. When the aggregation effect was significant, the RIs were determined using the subgroup of blood specimens with no or little aggregation. The effects of sex, age and weight were also investigated, but were moderate. The different RIs obtained with the Sysmex XT-2000iV and ProCyte DX analysers, and the two anticoagulants, were very similar to previous RIs established in EDTA with the ADVIA 120, another laser-based analyser, except for the platelet count in CTAD specimens. Its lower reference limit was higher in CTAD vs EDTA specimens, which confirms the interest in this anticoagulant in cats.

Changes in haematology measurements with the Sysmex XT-2000iV during storage of feline blood sampled in EDTA or EDTA plus CTAD.

In veterinary medicine a complete blood cell count (CBC) cannot always be performed within 24 h as usually recommended, particularly for specimens shipped to a reference laboratory. This raises the question of the stability of the variables, especially in ethylenediamine tetra-acetic acid (EDTA) feline blood specimens, known to be prone to in vitro platelet aggregation. Citrate, theophylline, adenosine and dipyridamole (CTAD) has been reported to limit platelet aggregation in feline blood specimens. The aim of this study was to measure the stability of the haematological variables and the platelet aggregation score in EDTA and EDTA plus CTAD (EDCT) feline blood specimens during 48 h of storage at room temperature. Forty-six feline EDTA and EDCT blood specimens were analysed with a Sysmex XT-2000iV analyser, and the platelet count and score of platelet aggregation were estimated immediately and after 24 and 48 h of storage. A significant increase in mean corpuscular volume, haematocrit, reticulocyte and eosinophil counts, and a significant decrease in mean corpuscular haemoglobin concentration and monocyte count were observed. Haemoglobin, mean corpuscular haemoglobin, and red blood cell, white blood cell, neutrophil and lymphocyte counts remained stable. Changes in reticulocyte indexes with time (low fluorescence ratio, medium fluorescence ratio, high fluorescence ratio and immature reticulocyte fraction) were not significant. Changes were generally more pronounced in EDTA than in EDCT. Platelet aggregation decreased markedly in initially highly aggregated EDTA specimens, and increased slightly in initially non- or mildly-aggregated EDTA or EDCT specimens. Platelet counts increased and decreased, or remained stable, respectively. CTAD can reduce storage-induced changes of the haematological variables in feline samples, thus improving the reliability of a CBC and limiting clinical misinterpretations.

Effects of intravenous, low-dose ketamine-diazepam sedation on the results of hematologic, plasma biochemical, and coagulation analyses in cats.

OBJECTIVE: To evaluate the effects of an IV, low-dose ketamine-diazepam combination used for short-duration chemical restraint on the results of clinicopathologic testing in cats and to assess its practicality and tolerance. DESIGN: Prospective case series. ANIMALS: 42 client-owned cats of various breeds, ages, and health status. PROCEDURES: Blood samples were obtained just prior to and just after IV injection of ketamine chlorhydrate (10 mg) and diazepam (0.5 mg). A CBC, plasma biochemistry panel, and coagulation profile were performed on each sample (ie, before and after chemical restraint). Practicality of the procedure was assessed, and cats were monitored for immediate and delayed effects. RESULTS: Significant changes were observed for most of the analytes tested. However, the magnitude of the observed changes was notably low and likely not of clinical relevance. The chemical-restraint procedure appeared effective, safe, and well tolerated. CONCLUSIONS AND CLINICAL RELEVANCE: The IV, low-dose ketamine-diazepam combination used for short-duration chemical restraint in the present study may be suitable to assist physical restraint for blood sampling for assessment of hematologic, serum biochemical, and coagulation parameters in cats.

Comparison of platelet clumping and complete blood count results with Sysmex XT-2000iV in feline blood sampled on EDTA or EDTA plus CTAD (citrate, theophylline, adenosine and dipyridamole).

False thrombocytopenia may result from platelet aggregation, especially in feline ethylenediamine tetra-acetic acid (EDTA) blood specimens. Citrate, theophylline, adenosine and dipyridamole (CTAD) was added to 46 feline EDTA specimens to test its anti-aggregation action. Platelet aggregation was estimated from blood films and a complete blood count was performed with a Sysmex XT-2000iV analyser. Platelet aggregation score was >2 in 11/46 EDTA tubes and only in one EDTA+CTAD specimen. The platelet count was higher in all CTAD-supplemented tubes except one, medians measured by cytometry being 225.5 × 10(9)/l and 249.0 × 10(9)/l in EDTA and EDTA+CTAD, respectively (P = 0.007). Adding CTAD had statistically and analytically significant but moderate effects on other blood variables, the most intense variations being observed for reticulocytes (about 3% higher in EDTA specimens) and reticulocyte indexes. Addition of CTAD to EDTA when sampling feline blood is a useful option to reduce platelet clumping.

Canine reference intervals for the Sysmex XT-2000iV hematology analyzer.

BACKGROUND: The laser-based Sysmex XT-2000iV hematology analyzer is increasingly used in veterinary clinical pathology laboratories, and instrument-specific reference intervals for dogs are not available. OBJECTIVE: The purpose of this study was to establish canine hematologic reference intervals according to International Federation of Clinical Chemistry and Clinical and Laboratory Standards Institute guidelines using the Sysmex XT-2000iV hematology analyzer. METHODS: Blood samples from 132 healthy purebred dogs from France, selected to represent the most prevalent canine breeds in France, were analyzed. Blood smears were scored for platelet (PLT) aggregates. Reference intervals were established using the nonparametric method. PLT and RBC counts obtained by impedance and optical methods were compared. Effects of sex and age on reference intervals were determined. RESULTS: The correlation between impedance (I) and optical (O) measurements of RBC and PLT counts was excellent (Pearson r=.99 and .98, respectively); however, there were significant differences between the 2 methods (Student's paired t-test, P<.0001). Differences between sexes were not significant except for HCT, PLT-I, and PLT-O. WBC, lymphocyte, and neutrophil counts decreased significantly with age (ANOVA, P<.05). Median eosinophil counts were higher in Brittany Spaniels (1.87 × 10(9) /L), Rottweilers (1.41 × 10(9) /L), and German Shepherd dogs (1.38 × 10(9) /L) than in the overall population (0.9 × 10(9) /L). PLT aggregates were responsible for lower PLT counts by the impedance, but not the optical, method. CONCLUSION: Reference intervals for hematologic analytes and indices were determined under controlled preanalytical and analytical conditions for a well-characterized population of dogs according to international recommendations.

Validation of preexisting reference intervals: can the procedure be applied to canine hemostasis?

The de novo establishment of reference intervals (RIs) for all variables is beyond the capabilities of many small laboratories. Thus, recent international recommendations propose procedures to adopt RIs established by "donor" laboratories after validation in "receiving" laboratories. The objective of the current study was to use recently published RIs of canine hemostasis tests as possible donor values and evaluate the validation procedure with randomized sets of values obtained in another study of canine RI determination of prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen, and antithrombin (AT). The preanalytical, analytical, and demographic conditions of the donor and receiving laboratories were first compared. To represent new reference individuals, 25 validation sample sets of 20 results of the receiving laboratory were randomly selected for each variable and compared with the RI of the donor laboratory. Validation was rejected in all cases for APTT and AT. Donor RI could be validated in 14 of 25 cases for fibrinogen and in 4 of 25 cases for PT. When preanalytical and analytical differences existed between donor and receiving laboratories, validation procedures consistently rejected preexisting RI. When the differences are smaller, the variability of the results obtained in the validation sample sets tested may be responsible for validations or rejections, which can lead to further misinterpretations of results from patients. Validation of a preexisting reference interval is certainly an interesting option for small laboratories, but progressive determination of the laboratory's own reference interval is probably a better long-term solution.