scholarly journals Diagnosis of Bovine Paratuberculosis by a Novel Enzyme-Linked Immunosorbent Assay Based on Early Secreted Antigens of Mycobacterium avium subsp. paratuberculosis

2008 ◽  
Vol 15 (8) ◽  
pp. 1277-1281 ◽  
Author(s):  
Sung Jae Shin ◽  
Donghee Cho ◽  
Michael T. Collins
Keyword(s):  
Immunosorbent Assay ◽  
Mycobacterium Avium ◽  
Area Under The Curve ◽  
Cost Effective ◽  
Enzyme Linked Immunosorbent Assay ◽  
Protein Antigens ◽  
Characteristic Analysis ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Cattle Herds ◽  
Low Passage

ABSTRACT We previously reported that protein antigens of serodiagnostic potential were more abundant in culture filtrates than cellular extracts from liquid cultures of Mycobacterium avium subsp. paratuberculosis (D. Cho and M. T. Collins, Clin. Vaccine Immunol. 13:1155-1161, 2006). Based on this observation, a novel enzyme-linked immunosorbent assay (ELISA) using antigens secreted by young (early- to mid-log-phase) cultures of M. avium subsp. paratuberculosis JTC303 (a low-passage isolate originating from the ileum of a Holstein bull) in mycobactin-supplemented Watson-Reid medium (pH 6.0) was developed and evaluated using a previously described panel of bovine sera (M. T. Collins et al., Clin. Diagn. Lab. Immunol. 12:685-692, 2005) that included 444 paratuberculosis cases and 412 controls. The new assay, called JTC-ELISA, had a significantly higher diagnostic sensitivity and an equivalent specificity compared to those of five commercial paratuberculosis ELISA kits. By receiver-operating characteristic analysis, the JTC-ELISA had the highest area under the curve of the six assays evaluated. The JTC-ELISA was particularly sensitive at detecting low-level fecal shedders of Mavium subsp. paratuberculosis (40%; the sensitivity of the commercial kits was 20%). The JTC-ELISA works effectively on both serum and milk samples for the detection of cattle with subclinical M. avium subsp. paratuberculosis infections, providing a cost-effective diagnostic tool to support paratuberculosis control programs in cattle herds.

scholarly journals Sensitive and Specific Enzyme-Linked Immunosorbent Assay for Detecting Serum Antibodies against Mycobacterium avium subsp. paratuberculosis in Fallow Deer

2014 ◽  
Vol 21 (8) ◽  
pp. 1077-1085 ◽  
Author(s):  
José M. Prieto ◽  
Ana Balseiro ◽  
Rosa Casais ◽  
Naiara Abendaño ◽  
Liam E. Fitzgerald ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Mycobacterium Avium ◽  
Cost Effective ◽  
Fallow Deer ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Igg Antibody ◽  
Tissue Samples ◽  
Content Type ◽  
Mycobacterium Avium Subsp Paratuberculosis

ABSTRACTThe enzyme-linked immunosorbent assay (ELISA) is the diagnostic test most commonly used in efforts to control paratuberculosis in domestic ruminants. However, commercial ELISAs have not been validated for detecting antibodies againstMycobacterium aviumsubsp.paratuberculosisin wild animals. In this study, we compared the sensitivities and specificities of five ELISAs using individual serum samples collected from 41 fallow deer with or without histopathological lesions consistent with paratuberculosis. Two target antigenic preparations were selected, an ethanol-treated protoplasmic preparation obtained from a fallow deerM. aviumsubsp.paratuberculosisisolate (ELISAs A and B) and a paratuberculosis protoplasmic antigen (PPA3) (ELISAs C and D). Fallow deer antibodies bound to the immobilized antigens were detected by using a horseradish peroxidase (HRP)-conjugated anti-fallow deer IgG antibody (ELISAs A and C) or HRP-conjugated protein G (ELISAs B and D). A commercially available assay, ELISA-E, which was designed to detectM. aviumsubsp.paratuberculosisantibodies in cattle, sheep, and goats, was also tested. Although ELISAs A, C, and E had the same sensitivity (72%), ELISAs A and C were more specific (100%) for detecting fallow deer with lesions consistent with paratuberculosis at necropsy than was the ELISA-E (87.5%). In addition, the ELISA-A was particularly sensitive for detecting fallow deer in the latent stages of infection (62.5%). The antibody responses detected with the ELISA-A correlated with both the severity of enteric lesions and the presence of acid-fast bacteria in gut tissue samples. In summary, our study shows that the ELISA-A can be a cost-effective diagnostic tool for preventing the spread of paratuberculosis among fallow deer populations.

scholarly journals Improved Sensitivity of Diagnosis of Tuberculosis in Patients in Korea via a Cocktail Enzyme-Linked Immunosorbent Assay Containing the Abundantly Expressed Antigens of the K Strain of Mycobacterium tuberculosis

2008 ◽  
Vol 15 (12) ◽  
pp. 1788-1795 ◽  
Author(s):  
A-Rum Shin ◽  
Sung Jae Shin ◽  
Kil-Soo Lee ◽  
Sun-Ho Eom ◽  
Seung-Sub Lee ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Immunosorbent Assay ◽  
Area Under The Curve ◽  
Infectious Agent ◽  
Enzyme Linked Immunosorbent Assay ◽  
Characteristic Analysis ◽  
Ionization Time ◽  
Flight Mass Spectrometry ◽  
Culture Filtrates ◽  
Single Antigen

ABSTRACT Tuberculosis (TB) is the leading cause of death from a single infectious agent in Korea. In this study, we compared the proteins present in culture filtrates from Mycobacterium tuberculosis strain K, which is the dominant clinical isolate in Korea, with those present in culture filtrates from M. tuberculosis H37Rv. Several differences in expression were detected between the two strains for those proteins with a molecular mass of <20 kDa. ESAT-6, HSP-X, and CFP-10 were found to be abundantly expressed in the strain K culture filtrates by liquid chromatography-electrospray ionization-time of flight mass spectrometry. The serodiagnostic potentials of recombinant antigens rESAT-6, rHSP-X, and rCFP-10 and two native antigens (Ag85 and PstS1) were evaluated by Western blot analysis and enzyme-linked immunosorbent assay (ELISA) using sera collected from 46 TB patients with active disease and 46 healthy controls. As for our ELISA results, HSP-X was superior to the other antigens in terms of sensitivity when a single antigen was employed. The results of a receiver operator characteristic analysis revealed that a cocktail ELISA using all five antigens was significantly more sensitive (77.8%) than the use of a single antigen and offered equivalent specificity; moreover, it produced the largest area under the curve (0.91 versus 0.55 to 0.87). Therefore, a cocktail ELISA containing abundantly expressed antigens enhances the sensitivity of a single antigen and can be a useful diagnostic tool for the detection of active TB.

scholarly journals Immunoglobulin G1 Enzyme-Linked Immunosorbent Assay for Diagnosis of Johne's Disease in Red Deer (Cervus elaphus)

2005 ◽  
Vol 12 (12) ◽  
pp. 1401-1409 ◽  
Author(s):  
J. Frank T. Griffin ◽  
Evelyn Spittle ◽  
Christie R. Rodgers ◽  
Simon Liggett ◽  
Marc Cooper ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Cervus Elaphus ◽  
Johne's Disease ◽  
Johne’S Disease ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Marker ◽  
Protein Antigens ◽  
Characteristic Analysis ◽  
Purified Protein Derivative ◽  
Subclinical Disease

ABSTRACT This study was designed to develop a customized enzyme-linked immunosorbent assay (ELISA) for the serodiagnosis of Johne's disease (JD) in farmed deer. Two antigens were selected on the basis of their superior diagnostic readouts: denatured purified protein derivative (PPDj) and undenatured protoplasmic antigen (PpAg). ELISA development was based on the antigen reactivity of the immunoglobulin G1 (IgG1) isotype, which is a highly specific marker for mycobacterial disease seroreactivity in deer. Sensitivity estimates and test parameters were established using 102 Mycobacterium paratuberculosis-infected animals from more than 10 deer herds, and specificity estimates were determined using 508 uninfected animals from 5 known disease-free herds. A receiver-operated characteristic analysis determined that at a cut point of 50 ELISA units, there was a specificity of 99.5% and sensitivities of 84.0% with PPDj antigen, 88.0% with PpAg, and 91.0% when the antigens were used serially in a composite test. Estimated sensitivity was further improved using recombinant protein antigens unique for M. paratuberculosis, which identified infected animals that were unreactive to PPDj or PpAg. While 80% of animals that were seropositive in the IgG1 ELISA had detectable histopathology, the assay could also detect animals with subclinical disease. The test was significantly less sensitive (75%) for animals that were culture positive for M. paratuberculosis but with no detectable pathology than for those with pathological evidence of JD (>90%). When the IgG1 ELISA was used annually over a 4-year period in a deer herd with high levels of clinical JD, it eliminated clinical disease, increased production levels, and reduced JD-related mortality.

scholarly journals Validation of a Monoclonal Antibody-Based Capture Enzyme-Linked Immunosorbent Assay for Detection of Campylobacter fetus

2005 ◽  
Vol 12 (11) ◽  
pp. 1261-1268 ◽  
Author(s):  
J. Devenish ◽  
B. Brooks ◽  
K. Perry ◽  
D. Milnes ◽  
T. Burke ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Immunosorbent Assay ◽  
Area Under The Curve ◽  
Stomach Contents ◽  
Enzyme Linked Immunosorbent Assay ◽  
Characteristic Analysis ◽  
Serotype A ◽  
Campylobacter Fetus ◽  
Field Samples ◽  
Serotype B

ABSTRACT A monoclonal antibody (MAb)-based antigen capture enzyme-linked immunosorbent assay (ELISA) was compared with the routine culture methodology for the detection of Campylobacter fetus subspecies from bovine and ovine field samples inoculated into Clark's transport enrichment medium (TEM). The work was a collaboration between two different diagnostic laboratories, one in Canada and the other in England. In both labs, TEM samples were incubated for 4 days at 35°C and then tested by culture and ELISA. The ELISA consisted of initial screening with MAb M1825 against C. fetus subspecies core lipopolysaccharide (LPS). All samples positive on ELISA screening were then retested by ELISA with MAb M1825 and MAbs M1177, M1183, and M1194, which recognize serotype A- and/or serotype B-specific C. fetus subspecies LPS epitopes. The Canadian samples consisted of 1,060 preputial washings from 529 bulls, of which 18 were positive by both culture and ELISA and 1,042 were negative by both methods. The English samples consisted of 321 tissue specimens, mostly stomach contents and placentas, from 190 aborted ovine and bovine fetuses. A total of 262 samples were negative by culture and ELISA, 52 samples were positive by culture and ELISA, and 7 samples were culture negative but ELISA positive. The results for all 70 culture-positive isolates were confirmed by conventional biochemical methods as C. fetus subsp. fetus, with 39 presumptively identified by the ELISA as serotype A and 30 presumptively identified as serotype B and with one sample containing isolates presumptively identified as serotype A and serotype B. A receiver operating characteristic analysis of the combined ELISA data from both countries resulted in an area under the curve of 0.997, with a sensitivity of 100% and a specificity of 99.5% relative to the results of culture. The data confirm that this ELISA method can be used as an excellent test for the screening of field samples in TEM for the presence of C. fetus subspecies.

scholarly journals Relationship between Presence of Cows with Milk Positive for Mycobacterium avium subsp. paratuberculosis-Specific Antibody by Enzyme-Linked Immunosorbent Assay and Viable M. avium subsp. paratuberculosis in Dust in Cattle Barns

2013 ◽  
Vol 79 (18) ◽  
pp. 5458-5464 ◽  
Author(s):  
Susanne W. F. Eisenberg ◽  
Ruj Chuchaisangrat ◽  
Mirjam Nielen ◽  
Ad P. Koets
Keyword(s):  
Dairy Cattle ◽  
Immunosorbent Assay ◽  
Mycobacterium Avium ◽  
Specific Antibody ◽  
Enzyme Linked Immunosorbent Assay ◽  
Young Stock ◽  
Content Type ◽  
Lactating Cows ◽  
Settled Dust ◽  
Mycobacterium Avium Subsp Paratuberculosis

ABSTRACTParatuberculosis, or Johne's disease, in cattle is caused byMycobacterium aviumsubsp.paratuberculosis, which has recently been suspected to be transmitted through dust. This longitudinal study on eight commercialM. aviumsubsp.paratuberculosis-positive dairy farms studied the relationship between the number of cows withM. aviumsubsp.paratuberculosisantibody-positive milk and the presence of viableM. aviumsubsp.paratuberculosisin settled-dust samples, including their temporal relationship. Milk and dust samples were collected in parallel monthly for 2 years.M. aviumsubsp.paratuberculosisantibodies in milk were measured by enzyme-linked immunosorbent assay (ELISA) and used as a proxy forM. aviumsubsp.paratuberculosisshedding. Settled-dust samples were collected by using electrostatic dust collectors (EDCs) at six locations in housing for dairy cattle and young stock. The presence of viableM. aviumsubsp.paratuberculosiswas identified by liquid culture and PCR. The results showed a positive relationship (odds ratio [OR], 1.2) between the number of cows with ELISA-positive milk and the odds of having positive EDCs in the same airspace as the adult dairy cattle. Moreover, the total number of lactating cows also showed an OR slightly above 1. This relationship remained the same for settled-dust samples collected up to 2 months before or after the time of milk sampling. The results suggest that removal of adult cows with milk positive forM. aviumsubsp.paratuberculosis-specific antibody by ELISA might result in a decrease in the presence of viableM. aviumsubsp.paratuberculosisin dust and therefore in the environment. However, this decrease is likely delayed by several weeks at least. In addition, the data support the notion thatM. aviumsubsp.paratuberculosisexposure of young stock is reduced by separate housing.

scholarly journals Sensitive Blood-Based Detection of Asbestos-Associated Diseases Using Cysteine-Rich Angiogenic Inducer 61 as Circulating Protein Biomarker

2020 ◽  
Author(s):  
Kai Bartkowiak ◽  
Swaantje Casjens ◽  
Antje Andreas ◽  
Lucija Ačkar ◽  
Simon A Joosse ◽  
...  
Keyword(s):  
Large Scale ◽  
Mononuclear Cells ◽  
Area Under The Curve ◽  
Immunofluorescent Staining ◽  
Youden Index ◽  
Enzyme Linked Immunosorbent Assay ◽  
Characteristic Analysis ◽  
Clinical Sensitivity ◽  
Associated Diseases ◽  
Healthy Control

Abstract Background Detection of asbestos-associated diseases like asbestosis or mesothelioma is still challenging. We sought to improve the diagnosis of benign asbestos-associated disease (BAAD) by detection of the protein cysteine-rich angiogenic inducer 61 (Cyr61) in human plasma. Methods Plasma Cyr61 was quantified using an enzyme-linked immunosorbent assay. Plasma samples from males diagnosed with BAAD, but without a malignant disease (n = 101), and malignant mesothelioma (n = 21; 15 males, 6 females), as well as nonasbestos-exposed healthy control participants (n = 150; 58 males, 92 females) were analyzed. Clinical sensitivity and specificity of Cyr61 were determined by receiver operating characteristic analysis. Results The median plasma Cyr61 concentration for healthy control participants was 0.27 ng/mL. Cytoplasmic Cyr61 in peripheral blood mononuclear cells from healthy control participants was evenly distributed, as detected by immunofluorescent staining. The increase in plasma Cyr61 concentrations in the BAAD study group was statistically significant compared to the healthy control participants (P &lt; 0.0001). For the detection of BAAD vs male healthy control participants, clinical sensitivity was 88% and clinical specificity 95% with an area under the curve of 0.924 at maximal Youden Index. For a predefined clinical specificity of 100%, the clinical sensitivity was 76%. For male mesothelioma patients vs male healthy control participants, the clinical sensitivity at maximal Youden Index was 95% with a clinical specificity of 100% (area under the curve, 0.997) and for a predefined clinical specificity of 100%, the clinical sensitivity was 93%. Conclusions In our study, plasma Cyr61 protein concentrations showed to be a new biomarker for asbestos-associated diseases like BAAD and mesothelioma in men, which deserves further investigation in large-scale cohort studies.

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