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SUMMARY

Five cats were made anemic by one-time phlebotomy, and their reticulocyte responses were monitored daily for 20 days, using manual enumeration and a standardized feline reticulocyte protocol developed and validated in our laboratory. The reticulocyte responses of 38 clinically normal client-owned cats also were analyzed manually and cytometrically to determine clinical reference ranges.

Increases in the percentage of aggregate reticulocytes over the reference range were detected in 5 of 5 phlebotomized cats, using the cytometric protocol. Only 4 of the 5 cats had an increase by results of manual enumeration. Manual aggregate counts had considerable daily variation and often fluctuated in and out of reference range, whereas cytometric aggregate counts remained consistently increased for distinct periods. Increased numbers of aggregate cells could also be detected for longer periods when evaluated by flow cytometry.

Increased numbers of punctate reticulocytes were detected in 4 of 5 cats, using the cytometric protocol. None of the cats had increased numbers of punctate cells when evaluated by use of the manual technique.

Aggregate reticulocytes in the 38 clinically normal cats ranged from 0.1 to 0.5%, which corresponded to 8,487 to 42,120 cells/μl. Punctate reticulocytes ranged from 2 to 17%, which corresponded to 225,400 to 1,268,584 cells/μl.

Flow cytometry, using a standardized analysis protocol, was a more reliable and sensitive technique for detection and evaluation of feline reticulocytosis than was manual enumeration. The sensitivity of the flow cytometer to small amounts of intracellular nucleoprotein makes this assay especially valuable for detection of punctate reticulocytosis and low degrees of aggregate reticulocytosis in cats.

Free access
in American Journal of Veterinary Research

Abstract

Objective

To evaluate a method for detecting thiazole orange-positive (TO+, reticulated) platelets in equine blood, using flow cytometry.

Animals

16 healthy, equine infectious anemia virus (EIAV)-negative horses and ponies; 9 thrombocytopenic, ElAV-positive horses and ponies; and 2 thrombocytopenic, ElAV-negative horses.

Procedure

Blood from healthy and thrombocytopenic horses was collected by jugular venipuncture. Appropriate sample requirement and incubation time for the assay were evaluated, using blood anticoagulated with EDTA or sodium citrate, or platelet-rich plasma in sodium citrate. The sample of blood or platelet-rich plasma was incubated with thiazole orange, and flow cytometric analysis was performed. Percentage of circulating TO+ platelets was determined from fluorescence (FL-1) logarithmic histograms.

Results

Healthy ponies (n = 9) had 1.28 to 2.83% (mean ± SD, 2.03 ± 0.50%) and horses (n = 7) had 0.9 to 3.44% (2.12 ± 1.14%) TO+ platelets in circulation. Thrombocytopenic ponies (n = 7) had 11.14 to 48.41 % (26.51 ± 11.99%) and thrombocytopenic horses (n = 4) had 2.33 to 8.52% (6.19 ± 2.68%) TO+ platelets in circulation. Mean platelet counts for the thrombocytopenic ponies and horses were 24,400 ± 20,500 and 39,300 ± 13,500 platelets/μl, respectively (reference range, 94,000 to 232,000 platelets/μl).

Conclusion

Thiazole orange-positive platelets can be detected in equine blood and percentages of TO+ platelets are increased in thrombocytopenic horses.

Clinical Relevance

Enumeration of TO+ platelets may prove to be a helpful noninvasive clinical measurement of bone marrow platelet production and aid in the assessment of platelet kinetics in thrombocytopenic horses. (Am J Vet Res 1997;58:1092–1096)

Free access
in American Journal of Veterinary Research