Antigen and antibody testing for the diagnosis of blastomycosis in dogs

BACKGROUND: Early diagnosis and treatment are associated with an improved prognosis in blastomycosis. The diagnosis of blastomycosis may be missed by cytology, histopathology, culture, or serology. An enzyme immunoassay (EIA) for detection of Blastomyces dermatitidis galactomannan antigen in body fluids has been used for rapid diagnosis of blastomycosis in humans. HYPOTHESIS: Measurement of Blastomyces antigen in urine or serum by the MVista Blastomyces antigen EIA is more sensitive than measurement of anti-Blastomyces antibodies for diagnosis of blastomycosis in dogs. METHODS: Serum and urine samples from 46 dogs with confirmed blastomycosis were tested for Blastomyces antigen and serum was tested for anti-Blastomyces antibodies. RESULTS: The sensitivity for the detection of antigen in urine was 93.5% and it was 87.0% in serum. The sensitivity of antibody detection by agar gel immunodiffusion (AGID) was 17.4% and it was 76.1% by EIA. Antigen and antibody decreased during itraconazole treatment. CONCLUSIONS AND CLINICAL IMPORTANCE: Antigen detection is a more sensitive test for diagnosis of blastomycosis than antibody testing by AGID, the only commercially available method. Antigen concentrations decreased with treatment.     

Blastomycosis in nondomestic felids.

Blastomycosis was diagnosed in six nondomestic felids from eastern Tennessee, including two Asian lions (Panthera leo persicus), one African lion (Panthera leo), one Siberian tiger (Panthera tigris), one cheetah (Acinonyx jubatus), and one snow leopard (Panthera uncia). Clinical signs included lethargy, anorexia, weight loss, dyspnea, sneezing. ataxia, and paresis. Variable nonspecific changes included leukocytosis, monocytosis, moderate left shift of neutrophils, moderate hypercalcemia, hyperproteinemia, and hyperglobulinemia. Thoracic radiographs revealed interstitial and alveolar changes, consolidation or collapse of a lung lobe, bullae formation, and a pulmonary mass. Agar gel immunodiffusion (AGID) serology for Blastomyces dermatitidis was performed in five felids and was positive in three. The tiger had cerebral blastomycosis and was positive for AGID serologic tests of both cerebrospinal fluid and serum. One percutaneous lung aspirate in the snow leopard and one bronchial aspirate in an Asian lion demonstrated B. dermatitidis organisms. whereas tracheal wash samples and a nasal discharge were nondiagnostic in others. Treatment with itraconazole was attempted in four cats. The tiger improved before euthanasia, whereas the others did not survive beyond initial treatments. In four felids, B. dermatitidis was found in the lungs and tracheobronchial lymph nodes associated with a florid pyogranulomatous reaction; the tiger had a pyogranulomatous encephalomyelitis, and the cheetah had a single pulmonary granuloma. Thoracic radiography, cytologic examination of lung lesion aspirates, and B. dermatitidis AGID serology should be performed on clinically ill zoo felids in endemic areas to rule out blastomycosis.     

Utility of diagnostic tests for and medical treatment of pulmonary blastomycosis in dogs: 125 cases (1989-2006)

OBJECTIVE: To compare results of the most common diagnostic tests for pulmonary blastomycosis in dogs, identify factors associated with outcome, and determine response to various antifungal treatment protocols. DESIGN: Retrospective case series. ANIMALS: 125 dogs with pulmonary blastomycosis. PROCEDURES: Medical records were reviewed, and information was obtained regarding diagnostic methods, results of routine laboratory testing, and radiographic response to antifungal treatment. RESULTS: 79 dogs survived, 38 died, and 8 were euthanized. Transthoracic fine-needle aspiration and transtracheal lavage were the most common diagnostic methods. Results of an agar gel immunodiffusion test for antibodies against Blastomyces dermatitidis were negative in 12 of 24 (50%) dogs. Only 3 of 94 (3.2%) dogs in which cytologic or histologic examination or bacterial culture of pulmonary samples were performed had any evidence of concurrent bacterial infection. The half-time for radiographic resolution of pulmonary infiltrates did not vary significantly with antifungal treatment, and use of a loading dosage of itraconazole was not associated with significant improvements in outcome or time to disease resolution. Dogs that died had a higher number of band neutrophils at initial examination, compared with those that survived. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that the agar gel immunodiffusion test should not be used as the sole diagnostic test for pulmonary blastomycosis in dogs, that concurrent bacterial pneumonia was uncommon in dogs with pulmonary blastomycosis, and that the rate with which pulmonary infiltrates resolved did not vary significantly among antifungal treatments.     

Comparison of four serologic tests for the detection of antibodies to bovine leukemia virus

Four tests for detection of antibodies to bovine leukemia virus (BLV) were compared. The sera that were tested came from cattle in naturally infected commercial dairy herds, cattle that were infected under experimental conditions, and cattle in an isolated BLV-free herd. The tests that were compared included a radioimmunoprecipitation assay (RIA) with p24 antigen, a RIA with glycoprotein (gp) antigen, an agar-gel immunodiffusion (AGID) test with gp antigen, and a virus-neutralization (VN) test that was based on inhibition of BLV-induced syncytia in cell culture. Results of the 4 serologic tests agreed for 96.8% of the sera from cattle in commercial herds. The gp RIA detected the greatest number of positive sera (188); it was followed in turn by the p24 RIA (187), the VN test (183), and the AGID test (176). The gpd RIA titers of the 12 sera that gave negative AGID results were 175 or less. In RIA, the percentage of precipitation of labeled antigen by positive sera was almost always higher with gp antigen than with p24 antigen. Satisfactory sensitivity in the p24 RIA required the acceptance of a low level of antigen precipitation, 15%, as a positive test. In the gp RIA, however, almost all positive sera precipitated at least 50% of the labeled antigen. Nonspecific precipitation of antigen in the RIA by sera from BLV-free cattle ranged from 4% to 10%. Examination of sequential serum samples from 17 experimentally infected cattle showed that BLV antibody was first detected 2 to 8 weeks after inoculation. In 9 cattle, seroconversion was detected simultaneously by all of the tests. Results from the other 8 cattle indicated that seroconversion could be detected first by p24 RIA, followed by the gp RIA and the VN test. The longest interval between RIA seroconversion and AGID seroconversion was 10 days. Monthly tests of sera from 10 laboratory cattle that were infected by contact exposure showed that 7 animals seroconverted in all tests at the same time. Two cattle were positive first in RIA, but the next month they were also positive in the VN and AGID tests. One animal was positive in the RIA and the VN test for 2 months before antibody was detected by AGID.     

Cultural and histopathologic confirmation of canine blastomycosis diagnosed by an agar-gel immunodiffusion test

Sixteen sera from 17 dogs with blastomycosis produced a precipitin band identical with the diagnostically significant precipitin A band formed by a Blastomyces dermatitidis reference antiserum and a soluble B dermatitidis yeast-form antigen in the agar-gel immunodiffusion test (94% sensitivity). The other serum from a dog with histopathologic demonstration of B dermatitidis in pulmonic tissues produced an unrelated precipitin band. Sixteen of the 17 diagnoses made by the detection of precipitin A were confirmed by isolation and culture of B dermatitidis or by histopathologic demonstration of the pathogen. Three cases were confirmed by cultural isolation only, 10 by histopathologic demonstration only, and 3 by both. In three other dogs given amphotericin B, there were demonstrable changes in serum precipitin A reactions.     

Transtracheal aspiration in the diagnosis of pulmonary blastomycosis (17 cases: 2000-2005)

Blastomyces dermatitidis is a common etiologic agent of fungal pneumonia in dogs. Definitive diagnosis is based on cytologic demonstration of the organism in affected tissues. Fluid obtained through transtracheal aspiration has previously been reported to have a low diagnostic yield for B. dermatitidis organisms. This retrospective study identified B. dermatitidis organisms in 76% of samples when transtracheal aspiration was performed in 17 nonsedated dogs with pulmonary blastomycosis. Transtracheal aspiration is a noninvasive and simple procedure that should be considered as an early diagnostic test whenever blastomycosis is a differential diagnosis in dogs with pulmonary disease.     

Development and evaluation of an enzyme-linked immunosorbent assay for detection of bovine antibodies to epizootic hemorrhagic disease of deer viruses.

An indirect enzyme-linked immunosorbent assay (I.ELISA) is described for detection of bovine serum antibody to epizootic hemorrhagic diseases of deer virus (EHDV). Serum samples, at a dilution of 1:200, were incubated with group-specific EHDV antigens, pre-adsorbed to microtiter plates. Bound antibodies were detected by a murine monoclonal antibody to bovine immunoglobulin (Ig)G1 (heavy-chain specific) conjugated with horseradish peroxidase. The performance of the I.ELISA in detecting antibodies to EHDV in sequential serum samples from calves experimentally infected with serotypes 1,2,3 and 4 was evaluated. The I.ELISA detected EHDV antibodies from 14 days postinfection when seroconversion by the standard agar gel immunodiffusion (AGID) test was also evident. The group-specific antibodies to EHDV increased exponentially during the first two to four weeks postinfection and remained relatively stable for about 12 months in some calves. Unlike observations with the AGID test, no reaction was seen in the I.ELISA between blue-tongue virus (BTV) antigen and sera from calves given a single dose of EHDV. The performance of the I.ELISA and AGID were compared using 3,135 AGID negative bovine field sera from herds in Ontario, Alberta and British Columbia and 130 AGID positive samples collected from cattle in 1987 and 1988 during and after outbreaks of EHD in the Okanagan Valley, British Columbia. The specificity and sensitivity of the assay relative to the AGID test were 99.3% and 91.5% respectively, with an overall agreement of 99.0% between the tests.(ABSTRACT TRUNCATED AT 250 WORDS)     

An international comparison of different laboratory tests for the diagnosis of bovine leukosis: suggestions for international standardization

A collaborative study was conducted to compare the detection limit of different laboratory tests for antibodies against bovine leukemia virus (BLV). Serum and milk samples were tested in agar gel immunodiffusion (AGID), different modifications of indirect ELISA, blocking ELISA and ELISA procedures using monoclonal antibodies to BLV gp51 or BLV p24. The detection limit of reference serum E4 diluted 2-fold in negative serum gave a median value of 1:16 in AGID, indirect ELISA, and monoclonal ELISA p24, 1:128 in monoclonal ELISA gp51, and 1:1024 in blocking ELISA. The detection limit of a 4% immunoglobulin preparation of E4 diluted in negative milk showed median values of 1:800 in indirect ELISA, 1:1000 in monoclonal ELISA, and 1:2400 in blocking ELISA. None of the ELISA procedures could detect all the positive individual milk samples diluted 1:50. The AGID test is the official reference test for detection of antibodies against BLV. Reference serum E4 diluted 1:10 in negative serum must be scored positive in the AGID test. It is suggested that an international reference serum standard be established rather than an official recommendation of a particular ELISA test.     

Mycotic mastitis in three dogs due to Blastomyces dermatitidis

Canine blastomycosis is a common systemic fungal infection within the Ohio and Mississippi River valleys and typically presents as pneumonia, lymphadenitis, or endophthalmitis. This report describes three cases in which mammary tissue samples were submitted to the Department of Pathobiology, University of Tennessee, College of Veterinary Medicine with clinical suspicion of neoplasia or postpartum bacterial mastitis. Pyogranulomatous to granulomatous mastitis and dermatitis with intralesional yeast consistent with Blastomyces dermatitidis were diagnosed. Two of the three dogs also had lymph node and pulmonary involvement. Mycotic mastitis due to Blastomyces dermatitidis is rarely reported and blastomycosis should be considered in the differential diagnosis of dogs with mammary lesions from endemic areas.     

Development and validation of a radioimmunoassay for the measurement of canine pancreatic lipase immunoreactivity in serum of dogs

OBJECTIVE: To develop and validate a radioimmunoassay (RIA) for measuring canine pancreatic lipase immunoreactivity (cPLI) in serum obtained from dogs. SAMPLE POPULATION: Serum samples from 47 healthy dogs. PROCEDURES: Canine pancreatic lipase (cPL) was purified from pancreatic specimens of dogs. Antibodies against cPL were raised in rabbits and purified by use of affinity chromatography. A tracer was produced by iodination of cPL with 125I. An RIA was established and validated by determination of sensitivity, working range, dilutional parallelism, spiking recovery, and intra- and interassay variability. A reference range for cPLI in serum was established by use of the central 95th percentile for samples obtained from 47 healthy dogs. RESULTS: Sensitivity and upper limit of the working range were 0.88 and 863 microg/L, respectively. Observed-to-expected ratios for serial dilutions ranged from 84.9 to 116.5% for 4 samples. Observed-to-expected ratios for spiking recovery ranged from 82.8 to 128.6% for 4 samples. Coefficients of variation for intra-assay variability for 4 serum samples were 18.3, 4.2, 3.5, and 8.9%, whereas interassay coefficients of variation were 29.2, 6.2, 3.9, and 4.4%, respectively. The reference range was 4.4 to 276.1 microg/L. CONCLUSIONS AND CLINICAL RELEVANCE: We conclude that the RIA described is sensitive, linear, accurate, precise, and reproducible, with limited accuracy in the high end of the working range and limited precision and reproducibility in the low end of the working range. Additional studies are needed to evaluate whether this degree of accuracy, precision, and reproducibility will negatively impact clinical use of this assay.     

Clinical survey of antibodies against red blood cells in horses after homologous blood transfusion

Serum samples of 20 horses were evaluated for antibodies against RBC after homologous blood transfusion. Transfusion-associated antibodies against RBC were detected in 10 horses. Antibodies recognizing horse blood group antigens Aa, Ae, Db, and Dc were identified. Antibodies against Aa were found in all samples from Aa-negative horses that were transfused with Aa-positive RBC. Antibodies against Aa persisted for at least 1 year after transfusion. Antibodies against Ae were detected in 7 of 8 horses transfused with Ae-positive RBC. Initial appearance and persistence of antibodies against Ae differed among the horses; antibodies were initially detected 1 week to 154 weeks after transfusion and disappeared as early as 4 weeks after transfusion. Antibodies against Db or Dc were detected in less than or equal to 33% of the horses that lacked Db or Dc antigens and were transfused with Db- or Dc-positive RBC. Antibodies against Db and Dc were initially detected in sera later than were the A-system antibodies. Three mares with transfusion-associated antibodies subsequently produced healthy offspring. Two foals had RBC antigens corresponding to their dam's alloantibodies; maternal colostrum with antibodies against Aa was withheld from the Aa-positive foal. The Db-positive foal remained healthy after nursing the mare with serum antibodies against Db.     

Comparison of the commercial agar-gel immunodiffusion test and radioimmunoprecipitation assay for detection of antibodies to bovine leukemia virus

In a double-blind study, the commercial agar-gel immunodiffusion test (AGID) was compared with a radioimmunoprecipitation assay (RIA) performed with glycoprotein (gp) antigen for detection of antibodies to bovine leukemia virus. Of 240 sera tested, 115 were from adult cows and 125 were from precolostral calves. Most adult animals were tested within 1 week of parturition. Sera from 74 cattle were positive and sera from 166 cattle were negative by gp RIA. Sensitivity of the AGID, compared with the gp RIA, was 85.1% when the test was read at 48 hours and was 94.6% when read at 72 hours. Specificity increased from 92.2% at 48 hours to 96.4% at 72 hours. Reading the AGID again at 72 hours also clarified most reactions that were questionable at 48 hours due to a haze around the test serum well. Of 3 RIA-positive precolostral calf sera, 2 were AGID-negative and 1 had a questionable reaction by the AGID at 48 hours. Of 5 RIA-positive sera that were AGID-negative at 48 hours, 2 were precolostral calves and 3 were cows tested at parturition. Of 166 RIA-negative reactions, none was falsely positive by the AGID at 48 or at 72 hours.     

Serologic prevalence of Sarcocystis neurona, Toxoplasma gondii, and Neospora caninum in horses in Brazil.

OBJECTIVE: To determine serologic prevalence of Sarcocystis neurona, Toxoplasma gondii, and Neospora caninum in horses in Brazil. DESIGN: Prevalence survey. ANIMALS: 101 Thoroughbreds in Brazil. PROCEDURE: Blood samples were obtained from horses and tested for serum antibodies against S neurona by use of an immunoblot procedure with culture-derived S neurona merozoites as antigen, and for serum antibodies against T gondii and N caninum by use of a modified agglutination test with formalin-preserved tachyzoites and mercaptoethanol. RESULTS: Antibodies against S neurona and T gondii were detected in 36 and 16 of 101 horses, respectively. Cross-reactivity between antibodies against T gondii and S neurona was not detected. Antibodies against N caninum were not detected in any samples. CONCLUSIONS AND CLINICAL RELEVANCE: The high prevalence of antibodies against S neurona detected in clinically normal horses emphasizes the importance of examining CSF for antibodies when establishing a diagnosis of equine protozoal myeloencephalitis.     

Counterimmunoelectrophoresis (immunoelectroosmosis) and serum electrophoretic pattern in serologic diagnosis of canine blastomycosis

Counterimmunoelectrophoresis (CIEP) with blastomyces and histoplasma antigens was used in a serologic study of 181 dogs clinically suspected of having blastomycosis and of 8 dogs with confirmed blastomycosis or histoplasmosis. Thirteen of the 181 dogs, positive by CIEP, were euthanatized, and the diagnosis was confirmed by cultivation and/or microscopic detection of Blastomyces dermatitidis. Additional CIEP-positive dogs were confirmed by staining of aspirates collected in vivo. Radiographic support for the diagnosis was reported in 4 other dogs in which histoplasmosis was excluded by a negative CIEP with histoplasma antigen. The precipitating antibody may disappear during the course of the disease, as it did in 1 dog treated with amphotericin B, but not cured. This dog reverted from CIEP-positive to CIEP-negative within 17 months of treatment (with a weak reaction after 10 months of treatment). The CIEP-detectable antibody was present only in 1 dog without a confirmation by histopathologic findings or cultivation among 24 well-documented cases and 181 total tested sera. The CIEP was more sensitive and specific than was the gel-diffusion precipitin test, eliminated the problems of anticomplementarity that often affected the results of complement-fixation tests with canine sera, and served well in detecting dogs with blastomycosis. Electrophoretic pattern of sera from CIEP-positive dogs with blastomycosis showed a decrease in albumin and an increase in alpha 2- and often in beta- and gamma-globulins, with a substantial decrease of the albumin/globulin ratio.     

Comparison of histologic lesions of endophthalmitis induced by Blastomyces dermatitidis in untreated and treated dogs: 36 cases (1986-2001)

OBJECTIVE: To compare prevalence of organisms and histologic changes in eyes from dogs with blastomycosis that were either untreated or undergoing treatment with itraconazole. DESIGN: Retrospective study. ANIMALS: 36 dogs with endophthalmitis associated with blastomycosis. PROCEDURE: Signalment, results of ophthalmic examination, and duration of treatment with itraconazole were extracted from medical records. Histologic sections from eyes were examined for prevalence and viability (ie, budding) of fungal organisms. A scoring system was devised to assess the degree of inflammation. RESULTS: Clinically, all eyes were blind and had signs of severe endophthalmitis. Histologically, the type and degree of inflammation and prevalence of Blastomyces dermatitidis were not significantly different between dogs treated with itraconazole and untreated dogs or among groups of dogs treated for different time periods (4 to 14, 15 to 28, or 29 to 72 days). Replication of the organisms in vascular tissues as well as avascular spaces in the eyes was similar in treated and untreated dogs. Lens rupture was seen in 12 of 29 (41%) eyes. CONCLUSIONS AND CLINICAL RELEVANCE: Persistence of inflammation in eyes of dogs with naturally occurring blastomycosis is likely attributable to the continued presence of B. dermatitidis, regardless of the duration of treatment with itraconazole. Lens capsule rupture, a common and previously unreported histologic finding, may contribute to cataract formation and continued inflammation.     

Detection of Antinuclear Antibodies by Indirect Immunofluorescence in Dog Sera: Comparison of Rat Liver Tissue and Human Epithelial-2 Cells as Antigenic Substrate

Rat liver sections and a human epithelial cell line (HEp-2) were compared as substrates for the detection of antinuclear antibodies (ANA) in the serum of normal dogs and dogs with suspected autoimmune disease, using a standard indirect immunofluorescence (IIF) technique. Antibody reactivity against rat hepatocyte nuclei was frequently found at low serum dilutions in normal dog sera. Using rat liver sections, a minimum significant positive titer, allowing negativity in more than 95% of normal dog sera, was found to be 1/100. With this titer, ANA positivity could be verified in 64 of 112 (57%) reanalysed serum samples from dogs with suspected autoimmune disease, earlier determined as ANA-positive. No reactivity against nuclei of HEp-2 cells was observed in any of the normal dog sera analyzed at a screening dilution of 1/25. Using this dilution as a minimum significant positive titer, 63 of the 112 (56%) re-evaluated serum samples were positive. These 63 samples were from the same dogs as the 64 samples that were positive on rat liver sections. Thus, the 2 methods of ANA-IIF detected a nearly identical population of dogs with suspected autoimmune disease once the level of significance of a positive titer was adjusted to 195% specificity for each method. HEp-2 cells were found to be superior to rat liver cryostate sections as ANA substrate because of their low reactivity with normal sera, and the ease of discernment of the ANA fluorescence pattern. The recognition and documentation of specific pattern types may give clues to ANA subspecificities, which could prove useful if they are found to correlate with well-defined subgroups of immune mediated clinical diseases in dogs. J Vet Intern Med 1996;10:199–203. Copyright © 1996 by the American College of Veterinary Internal Medicine .     

Evaluation of serologic methods for the detection of antibodies to encephalomyocarditis virus in swine fetal thoracic fluids.

Hemagglutination inhibition (HI) and agar gel immunodiffusion (AGID) tests were compared to the serum neutralization (SN) test to evaluate their ability to detect antibodies to encephalomyocarditis virus (EMCV). Swine fetal thoracic fluids of known EMCV SN antibody titers (200 samples greater than or equal to 1:2, 100 samples less than 1:2) were selected from a collection of field cases. The thoracic fluids were tested for EMCV antibodies by HI and AGID, and the results were compared to those of the SN test. Of 200 SN antibody-positive samples, 183 (91.5%) and 173 (86.5%) were positive in HI and AGID tests, respectively. Of 100 SN-negative samples, 81 (81%) and 94 (94%) were negative in HI and AGID tests, respectively. Agreement between the tests was analyzed by calculating Kappa values. The values were 0.73 between SN and HI tests and 0.77 between SN and AGID tests, indicating very good to excellent agreement for HI and AGID tests with the SN test. Of 200 SN-positive samples, 19 samples with low SN titers (1:2-1:16) were further tested by Western immunoblotting, and all were confirmed as positive. Interpretation of the present results suggests that both HI and AGID tests can be used as alternatives to the SN test.     

Reactivity of serum samples of dogs and horses tested by use of class-specific recombinant-based enzyme-linked immunosorbent assays for detection of granulocytic ehrlichiosis

OBJECTIVE: To test serum samples of dogs and horses by use of class-specific recombinant-based ELISA for establishing a diagnosis of granulocytic ehrlichiosis attributable to infection with organisms from the Ehrlichia phagocytophila genogroup. SAMPLE POPULATION: Serum samples from 43 client-owned dogs and 131 horses (81 with signs of acute illness and 50 without signs of disease). PROCEDURE: Serum samples were analyzed, using ELISA with a recombinant 44-kd protein antigen for IgM and IgG antibodies to the human granulocytic ehrlichiosis (HGE) agent (NCH-1 strain). Western blot analyses, using infected human promyelocytic leukemia cells, were conducted on 38 serum samples of horses and 11 serum samples of dogs to verify reactivity to the 44-kd peptide. RESULTS: IgM or IgG antibodies to the HGE agent were detected in 5 to 28% of dog serum samples and 5 to 37% of horse serum samples. Thirty-five of 38 (92%) horse serum samples had corresponding results on both tests (2 positive results for 26 samples and 2 negative results for 9 samples), using an ELISA for IgG antibodies or immunoblotting for total immunoglobulins. All 11 serum samples of dogs had positive results for both methods. CONCLUSION AND CLINICAL RELEVANCE: These ELISA with recombinant 44-kd antigen are suitable for detecting IgM or IgG antibodies to the HGE agent in serum samples of dogs and horses. Positive results for serum samples of horses from Connecticut, New York, Virginia, and Georgia indicate that the HGE agent is widely distributed in tick-infested areas of the eastern United States.     

Seroprevalence of Neospora caninum in dogs in south-western Poland

The prevalence of anti-Neospora caninum antibodies was determined in serum samples from 110 dogs in south-western Poland, using the indirect fluorescent antibody test (IFAT). Antibodies to N. caninum were found in 18 dogs (16.36%), of which 7 were also positive to Toxoplasma gondii. The titres of anti-N. caninum antibodies varied from 1:50 to 1:200.     

Enzyme-linked immunosorbent assay for the diagnosis of bovine leukosis: comparison with the agar gel immunodiffusion test approved by the Canadian Food Inspection Agency.

Four commercially available bovine leukemia virus (BLV)-ELISA kits from Europe or the United States were compared to the agar gel immunodiffusion (AGID) test officially approved by the Canadian Food Inspection Agency (CFIA). A total of 1200 cattle serum samples were used. Three ELISA kits based on the envelope glycoprotein (gp51) gave an excellent correlation with the AGID test. The kappa values were 0.998, 0.984, and 0.986 for the ELISA kits #1, #2, and #3, respectively. The ELISA kit based on the p24 core protein was found to be less sensitive than the officially approved AGID test and detected 5.13% of false negatives. Forty BLV AGID strongly positive serum samples were diluted. Based on the dilution experiment, the gp51 ELISA kits were found to be more sensitive than the AGID test kits. They were capable of detecting antibodies in samples diluted up to 1/5000 (kit #1), 1/20 800 (kit #2) and 1/4000 (kit #3), whereas the AGID kit was only capable of detecting antibodies in samples diluted up to 1/100. Based on these observations, the gp51 BLV-ELISA was recognized as an official test method for the serodiagnosis of bovine leukosis in Canada.