Classes of antibody bound to erythrocytes were determined using direct immunofluorescence (DIF) flow cytometry in 3 horses and 12 dogs with immune-mediated hemolytic anemia (IMHA). Background levels of antibody binding were determined in samples from 12 horses and 12 dogs that were free of clinical disease. The range of nonspecific binding of a fluorescein isothiocyanate (FITC)-conjugated goat anti-equine immunoglobulin G (IgG) was 19.9-36.7%, but was eliminated by the use of the F(abЈ) 2 fragment of FITC-conjugated goat anti-equine IgG. Background binding by other class-specific antibodies to equine and canine erythrocytes was negligible. The DIF results were compared to the direct antiglobulin (Coombs') test in 5 horses and 20 dogs with anemia. The former assay was more sensitive in dogs with IMHA than was the Coombs' test (100% versus 58%). In contrast, the Coombs' test had better specificity than the DIF assay (100% versus 87.5%, respectively). Using clinical parameters or response to therapy as the comparison, the positive and negative predictive values for the DIF test were 92% and 100% compared to the values of the Coombs' test of 100% and 62%. The DIF assay detected low levels of cells bound with antibody (Ͻ30%) in 5 dogs that were Coombs' test-negative. For both species, performance of the DIF test was independent of the prozone effect. Five dogs with IMHA had IgG and IgM on erythrocytes, 5 had IgG, and 2 had IgM. Three horses had surface-bound IgG, including a horse with suspected penicillin-induced IMHA, a foal with neonatal isoerythrolysis, and a foal with clostridial septicemia. The DIF method was valuable in monitoring the response to therapy in the foal with neonatal isoerythrolysis.Key words: Antibody classes; Coombs' test; Direct immunofluorescence; Erythrocyte antibody; Flow cytometry. Immune-mediated hemolytic anemia (IMHA) is an immunohematologic disorder in which destruction of red blood cells is accelerated by the attachment of antibody, with or without complement, to the erythrocyte membrane. Antibodies may be directed against unaltered red blood cells (primary or idiopathic) or against erythrocytes that have been antigenically altered through interaction with secondary causes, including drugs, neoplasia, and infectious diseases.1-6 The relative frequencies of primary and secondary IMHA in dogs are 43% and 57%, respectively. 7The diagnosis is confirmed by the presence of spherocytosis, autoagglutination, or by a positive direct antiglobulin (Coombs') test that has been validated for use in the species of interest. [8][9][10][11][12] The Coombs' test is based on detection of agglutination or clumping of erythrocytes after addition of an anti-species polyvalent mixture of antibody to immunoglobulin M (IgM), IgG, and complement protein C 3 . Serial dilutions of the polyvalent Coombs' reagent are prepared and tested against patient erythrocytes to provide the proper concentration equivalence between antiglobulin and the antibody-coated erythrocytes at which agglutination occurs. Because of the...

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A 4-year-old Paint mare was examined because of respiratory tract infection, dermatitis, and weight loss of 2 months' duration. Initial examination revealed generalized pruritic dermatitis, ocular and nasal discharges, and stranguria. Laboratory abnormalities included leukopenia and hypoalbuminemia. Further examination of the respiratory tract revealed grade III of IV pharyngitis and pyogranulomatous pneumonia. Endoscopic examination of the bladder revealed a prolific mass at the junction of the bladder and urethra. Hypoproteinemia was suspected to be caused by protein-losing enteropathy. On histologic examination, skin, rectal, pharyngeal, and urethral biopsy specimens were characterized by infiltration of eosinophils and lymphocytes, and a diagnosis of multisystemic eosinophilic epitheliotropic disease was made. The horse improved following treatment with dexamethasone, trimethoprim-sulfamethoxazole, and an antihistamine and was discharged after 19 days of hospitalization. Treatment with dexamethasone was continued for 4 weeks after hospitalization but was then discontinued. Eight months after discharge, the horse was performing as a pleasure horse and did not require any medical treatment. Multisystemic eosinophilic epitheliotropic disease is typically associated with a poor prognosis in horses. The dermatitis, protein-losing enteropathy, and lower respiratory tract disease in this horse were consistent with previous reports; however, pharyngitis and urethritis have not, to our knowledge, been previously reported in horses with this disease.

A 9-year-old pregnant mare was referred for evaluation of a nonhealing wound of 8 weeks' duration on the lateral aspect of the left forelimb. A soft tissue mass encircled the proximal two thirds of the metacarpus; radiography revealed a moderate periosteal reaction affecting metacarpal bone i.v. Histologic and immunohistochemical examinations revealed eosinophilic granulomatous inflammation and Pythium sp in the soft tissues. The mare was treated for 12 days with antimicrobials, medicated wound dressings, debridement, and i.v. administration of sodium iodide; radiography revealed progression of the bone lesions. The mare was treated by regional arterial perfusion with miconazole and excision of affected soft tissues and the distal two thirds of metacarpal bone i.v. The mare recovered without complications and gave birth to a healthy foal. Regional perfusion of antifungal agents provides high concentrations in soft and osseous tissues and permits use of low dosages of agents administered by other routes, which reduces cost, adverse effects, and teratogenic effects.

Formalin was injected into an ethmoidal hematoma in an 18-year-old Arabian gelding. Abnormal neurologic signs were observed within minutes of the injection. The horse did not respond favorably to medical treatment of the neurologic signs and was euthanatized. Postmortem examination revealed erosion and necrosis of the ventral cribriform plate, which appeared to have allowed the injected formalin to reach the rostral portion of the frontal lobe of the brain. Endoscopy and radiography had been performed prior to euthanasia, but neither delineated the cribriform lesion. Before treating large progressive ethmoidal hematomas with formalin, it may be beneficial to perform computed tomography to assess the extent of damage caused by the lesion.