Application of a Direct Flow Cytometric Erythrocyte Immunofluorescence Assay in Dogs with Immune-Mediated Hemolytic Anemia and Comparison to the Direct Antiglobulin Test

Quigley, Kelley A.; Chelack, Brian J.; Haines, Deborah M.; Jackson, Marion L.

doi:10.1177/104063870101300403

Cited by 41 publications

(58 citation statements)

References 15 publications

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“…In some dogs with IMHA, the target antigens for RBC autoantibodies have been identified, although in most dogs with IMHA the antigens are unknown 8 . Previous studies have found that both immunoglobulin (Ig) G and IgM antibodies can be detected on the surface of RBC 7,9–13 . The primary consequence of antibody binding to the RBC surface is premature removal of RBC from circulation by macrophages in the spleen and liver 2–4 …”

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confidence: 99%

Anti‐Erythrocyte Antibodies and Disease Associations in Anemic and Nonanemic Dogs

Morley

Mathes

Guth

et al. 2008

Veterinary Internal Medicne

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Background: Flow cytometry has been used to detect anti-red blood cell (RBC) antibodies in dogs with immune-mediated hemolytic anemia (IMHA), but the prevalence of anti-RBC antibodies in anemic and nonanemic dogs with a variety of different diseases has not been assessed previously.Hypothesis: We hypothesized that anti-RBC antibodies would be more common in anemic dogs and in dogs with immunemediated disorders and cancer.Animals: Blood samples from 292 dogs were analyzed prospectively by flow cytometry for anti-RBC antibodies.Methods: Blood samples from 147 anemic and 145 nonanemic dogs were evaluated by flow cytometry to detect surfacebound immunoglobulin (Ig) G and IgM antibodies on RBC. Disease associations with RBC antibodies were determined, as was the correlation between disease status and the percentage of Ig 1 RBC. The specificity and sensitivity of flow cytometry and clinical variables for the diagnosis of IMHA were compared by Bayesian analysis.Results: Anemic dogs were significantly more likely to be positive for anti-RBC antibodies (IgG, IgM, or both) than nonanemic dogs. Anemic dogs also had significantly higher percentages of Ig 1 RBC than nonanemic dogs, whereas dogs with IMHA had significantly higher percentages of Ig 1 RBC than dogs with all other diseases. Dogs with IMHA, infectious diseases, and immune-mediated thrombocytopenia were significantly more likely to have anti-RBC antibodies than dogs with other medical or surgical diseases.Conclusions: Anemic dogs with immune-mediated diseases and infectious diseases were at the highest risk for the development of anti-RBC antibodies, and flow cytometry for the detection of IgG on RBC was highly sensitive and specific for the diagnosis of IMHA.

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confidence: 99%

Anti‐Erythrocyte Antibodies and Disease Associations in Anemic and Nonanemic Dogs

Morley

Mathes

Guth

et al. 2008

Veterinary Internal Medicne

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“…A anemia hemolítica imuno-mediada (AHIM) é a causa mais comum dentre as anemias hemolíticas e a doença imuno-mediada de maior prevalência em cães (Mackin 2000, Quigley et al 2001, McCullough 2003, Giger 2005. A AHIM é caracterizada pela destruição direta ou fagocitose de hemácias opsonizadas por imunoglobulinas como IgG ou IgM e/ou complemento.…”

Section: Introductionunclassified

Avaliação das alterações hemostáticas e do risco tromboembólico em cães com AHIM

et al. 2016

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ABSTRACT.-Moraes L.F., Takahira Immune-mediated hemolytic anemia (IMHA) is the most common cause of hemolytic anemia and the most prevalent immune-mediated disease in dogs, which is classified in primary or secondary. IMHA has been associated with hypercoagulability state and thromboembolism has been referred as the most common complication. The purpose of this study was to correlate the possibility of hemostatic abnormalities and the thromboembolic risk in IMHA and in anemias of other etiologies. 76 dogs were selected, all exhibiting hematocrit lower than 20% and at least one clinical sign commonly associated with IMHA or breed predisposition. These dogs were tested for IMHA by flow cytometry. Hemostatic tests (platelet count, PT, APTT, TT, AT, PDF and D-dimer) as well as CBC, reticulocyte count, blood parasite search in peripheral blood smear, Ehrlichia sp. and leptospirosis tests were performed in all dogs. 59 dogs were positive for IMHA. The thromboembolic risk was characterized by the presence of three or more changes in the tests of the hemostatic profile. 74.6% cases of IMHA were attributed to infectious diseases and was mostly associated with Ehrlichia sp. (88.6%). 72.1% of dogs had thrombocytopenia and 57.6% had regenerative anemia with significantly higher values of metarubricyte and reticulocyte count. There were no significant changes between two anemic groups (positive and negative for IMHA). Anemic dogs had higher APTT mean value and lower AT and platelet counts mean values than the control group (p<0.05). 25 dogs with IMHA and seven dogs without IHMA had thromboembolic state. The specificity of PDF was lower (30.2%) than previous studies. The choice of the Ig class does not affect the diagnosis of IMHA. There were no correlation between the presence of antibodies bound to the RBC surface and the hemostatic tests, but there were a week correlation (p<0,05) between Ht(%) and APTT (r=-0.2621), AT (r=0.4297) and platelets count (r=0.5349) values. Anemia is associated with hemostatic abnormalities and thromboembolic risk, however the immune-mediated nature of the disease was not the determining factor for this condition. In part, these changes can be attributed to infection diseases seen in both groups (positive and negative for IMHA). The similar thromboembolic risk evidenced in both groups of anemic dogs does not exclude the relationship of immune-mediated involvement, but alert to the possibility of thromboembolism in anemic dogs with Ehrlichia sp. (negative for IHMA). Therefore, anemic dogs with clinical signs associated with thromboembolic risk should be evaluated for this complication, today poorly 1 Recebido em 6 de junho de 2015.Aceito para publicação em 8 de março de 2016.

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“…Immunofluorescence Assay: The presence of IgG and/or IgM antibodies on the plasma membrane of erythrocytes and platelets was examined by flow cytometry according to the method described by Quigley et al (2001) on whole blood collected from all rats/group on week 5 (erythrocyte) and from the first 5 rats/group from the control, middose females, and high-dose males on week 10 (platelets). Antibodies used were the goat anti-rat IgG FITC (Bethyl Laboratories, Montgomery, TX) and mouse anti-rat IgM FITC (Clone HIS49; BD Bioscience, San Jose, CA).…”

Section: Direct Flow Cytometric Erythrocyte and Plateletmentioning

confidence: 99%

Drug-induced Hemolytic Anemia and Thrombocytopenia Associated with Alterations of Cell Membrane Lipids and Acanthocyte Formation

et al. 2010

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CXCR3 is a chemokine receptor, upregulated upon activation of T cells and expressed on nearly 100% of T cells in sites of inflammation. SCH 900875 is a selective CXCR3 receptor antagonist. Thrombocytopenia and severe hemolytic anemia with acanthocytosis occurred in rats at doses of 75, 100, and 150 mg/kg/day. Massively enlarged spleens corresponded histologically to extramedullary hematopoiesis, macrophages, and hemosiderin pigment and sinus congestion. Phagocytosed erythrocytes and platelets were within splenic macrophages. IgG and/or IgM were not detected on erythrocyte and platelet membranes. Ex vivo increased osmotic fragility of RBCs was observed. Lipid analysis of the RBC membrane revealed modifications in phosphatidylcholine, overall cholesterol, and/or sphingomyelin. Platelets exhibited slender filiform processes on their plasma membranes, analogous to those of acanthocytes. The presence of similar morphological abnormalities in acanthocytes and platelets suggests that possibly similar alterations in the lipid composition of the plasma membrane have taken place in both cell types. This phenotype correlated with alterations in plasma lipids (hypercholesterolemia and low triglycerides) that occurred after SCH 900875 administration, although other factors cannot be excluded. The increased cell destruction was considered triggered by alterations in the lipid profile of the plasma membranes of erythrocytes and platelets, as reflected morphologically.

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Application of a Direct Flow Cytometric Erythrocyte Immunofluorescence Assay in Dogs with Immune-Mediated Hemolytic Anemia and Comparison to the Direct Antiglobulin Test

Cited by 41 publications

References 15 publications

Anti‐Erythrocyte Antibodies and Disease Associations in Anemic and Nonanemic Dogs

Anti‐Erythrocyte Antibodies and Disease Associations in Anemic and Nonanemic Dogs

Avaliação das alterações hemostáticas e do risco tromboembólico em cães com AHIM

Drug-induced Hemolytic Anemia and Thrombocytopenia Associated with Alterations of Cell Membrane Lipids and Acanthocyte Formation

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