Development of an immunobinding dot-blot assay as an alternative for the serodiagnosis of human cysticercosis

2009 ◽  
Vol 83 (4) ◽  
pp. 333-337 ◽  
Author(s):  
E. Hernández-Cruz ◽  
J.J. González-Cabriales ◽  
C. Ordaz-Pichardo ◽  
N.I. de la Cruz-Hernández ◽  
G.H. Flores-Gutiérrez

AbstractAn immunobinding dot-blot assay (DBA) was developed on nitrocellulose paper for the serodiagnosis of human cysticercosis, using Cysticercus cellulosae as antigen. The DBA had an immunological sensitivity of 0.08 mg of antigen protein/ml; however, it showed cross-reactions with antigens of adult Taenia solium and Echinococcus granulosus, but not with Toxoplasma gondii and Entamoeba histolytica antigens. An enzyme-linked immunosorbent assay (ELISA) was used as the gold standard for obtaining the diagnostic validity of the DBA, giving 84.61%, 100.00%, 100.00% and 97.98% for epidemiological sensitivity, epidemiological specificity and positive and negative predictive values, respectively. There were no statistical differences between the two tests (P < 0.05, kappa = 0.907). This study showed that DBA is an alternative method for the serodiagnosis of human cysticercosis.

2018 ◽  
Vol 50 (3) ◽  
pp. 229-235 ◽  
Author(s):  
Suresh Bokoliya ◽  
Shripad Patil ◽  
Madhu Nagappa ◽  
Arun Taly

AbstractObjectiveTo assess the practicality of dot-blot testing for rapid and sensitive detection of the antiacetylcholine receptor (anti-AChR) antibodies in myasthenia gravis (MG).MethodsIn this case-control study, we tested serum specimens of 85 patients with MG, 85 healthy control individuals, and 85 patients without MG who have other autoimmune and neurological illnesses. All the serum specimens were tested for anti-AChR antibodies using 3 assays: in-house enzyme-linked immunosorbent assay (ELISA), the dot-blot assay, and commercial ELISA.ResultsIn-house ELISA, commercial ELISA, and dot-blot test results were positive for anti-AChR antibodies in 65 (76.5%) patients with MG. The results of all 3 tests were negative for anti-AChR antibodies in healthy controls and patients without MG. We observed perfect concordance (K = 1, P <.001) between all 3 tests. In-house ELISA correlated significantly (r = 0.873, P <.001) with commercial ELISA. In-house ELISA and the dot-blot test demonstrated similar diagnostic performance in detecting anti-AChR antibodies.ConclusionsThe dot-blot assay is a simple, nonradioactive immune assay for rapid detection of anti-AChR antibodies in MG.


2011 ◽  
Vol 24 (1) ◽  
pp. 42-50 ◽  
Author(s):  
Ming Yang ◽  
Rebekah van Bruggen ◽  
Wanhong Xu

Seneca Valley virus (SVV), a member of the Picornaviridae family, was implicated in a suspicious vesicular disease discovered in pigs from Canada in 2007. Because any outbreak of vesicular disease in pigs is assumed to be foot-and-mouth disease (FMD) until confirmed otherwise, a test for diagnosing the presence of SVV would be a very useful tool. To develop the diagnostic tests for SVV infection, 5 monoclonal antibodies (mAbs) were produced from mice immunized with binary ethylenimine (BEI)-inactivated SVV. Using a dot blot assay, the reactivity of the mAbs was confirmed to be specific for SVV, not reacting with any of the other vesicular disease viruses tested. The mAbs demonstrated reactivity with SVV antigen in infected cells by an immunohistochemistry assay. An SVV-specific competitive enzyme-linked immunosorbent assay (cELISA) was developed using BEI-inactivated SVV antigen and a mAb for serodiagnosis. The cELISA results were compared to the indirect isotype (immunoglobulin [Ig]M and IgG) ELISA and the virus neutralization test. All SVV experimentally inoculated pigs exhibited a positive SVV-specific antibody response at 6 days postinoculation, and the sera remained positive until the end of the experiment on day 57 (>40% inhibition) using the cELISA. The cELISA reflected the profile of the indirect ELISA for both IgM and IgG. This panel of SVV-specific mAbs is valuable for the identification of SVV antigen and the serological detection of SVV-specific antibodies.


2001 ◽  
Vol 8 (1) ◽  
pp. 79-84 ◽  
Author(s):  
Paul T. Fawcett ◽  
Carlos D. Rose ◽  
Sandra M. Budd ◽  
Kathleen M. Gibney

ABSTRACT This study evaluated the effects of vaccination with OspA on the use of serologic tests as aids in the diagnosis of Lyme borreliosis. Sera from control and OspA-immunized mice and from OspA-immunized human volunteers were tested for serologic reactivity to Borrelia burgdorferi. Testing was performed with samples obtained prior to administration of vaccine and at 30 days following administration of an initial and a second dose of OspA vaccine. The assays used to assess serologic reactivity included an in-house-developed enzyme-linked immunosorbent assay (ELISA), an in-house-developed Western blot assay, two commercial Western blot tests, and a commercially available dot blot assay. Data obtained from this study demonstrate that immunization with the OspA vaccine will cause ELISA to yield positive results (as reported previously) for the majority of vaccine recipients. Results obtained from Western blot analysis indicate that vaccination with recombinant OspA induces production of antibodies which bind to several different borrelial proteins. The degree of reactivity detected by Western blotting varied greatly between the three assays used. The in-house assay showed the least reactivity, while one commercial Western blot test actually yielded positive test results for infection with B. burgdorferi. The usefulness of all three Western blot assays for the diagnosis of potential infection in a vaccine recipient is severely limited by the extensive reactivity caused by vaccination alone. Antibodies produced in response to OspA vaccination did not significantly affect the performance of the dot blot test; thus, it could provide a reliable means to test for infection withB. burgdorferi in OspA vaccine recipients.


2010 ◽  
Vol 17 (9) ◽  
pp. 1398-1406 ◽  
Author(s):  
Ming Yang ◽  
Alfonso Clavijo ◽  
Jill Graham ◽  
John Pasick ◽  
James Neufeld ◽  
...  

ABSTRACT A panel of monoclonal antibodies (MAbs) was generated from mice immunized with binary ethylenimine (BEI)-inactivated H7N1 (A/TK/ON/18-2/00) virus. Using a dot blot assay, six of seven MAbs reacted with viruses of the H7 subtype, but not with any of the other 15 hemagglutinin (HA) subtypes tested. Four of the seven MAbs reacted with 14 different H7 isolates, indicating that the MAbs binding epitopes are conserved among viruses of the H7 subtype. The binding epitopes of all seven MAbs were conformational and reacted with the HA1 fraction of the HA protein in Western blots under nonreducing conditions. Applications of these MAbs in the development of rapid tests for H7 subtype viruses were evaluated. The MAbs demonstrated reactivity with AI virus H7 antigen in immunofluorescence and immunohistochemistry assays. Monoclonal antibody 3 showed a very strong immunostaining in the formalin-fixed and paraffin-embedded tissue from the H7N3 virus-infected chicken. A double-antibody sandwich (DAS) enzyme-linked immunosorbent assay (ELISA) was developed using two of the MAbs. The DAS ELISA specifically detected all H7 strains tested in this study. A competitive ELISA (cELISA) for the detection of H7-specific antibodies was evaluated using one MAb and BEI-inactivated H7N1 virus as the antigen. All infected birds showed positive antibody responses at 7 days postinfection. The sensitivity of this cELISA was comparable with that of an influenza A nucleoprotein-based cELISA. This panel of MAbs is valuable in the development of various immunoassays.


2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Mwemezi L. Kabululu ◽  
Maria V. Johansen ◽  
James E. D. Mlangwa ◽  
Ernatus M. Mkupasi ◽  
Uffe C. Braae ◽  
...  

Abstract Background Taenia solium is a zoonotic parasite responsible for neurocysticercosis—a major cause of late-onset acquired epilepsy in humans. Lack of affordable, specific and sensitive diagnostic tools hampers control of the parasite. This study assessed the performance of an antigen detection enzyme-linked immunosorbent assay (Ag-ELISA) in the diagnosis of viable T. solium cysticercosis in naturally infected slaughter-age pigs in an endemic area in Tanzania. Methods A total of 350 pigs were bled before they were slaughtered and their carcases examined. Serum was analyzed for circulating antigens by using a monoclonal antibody-based B158/B60 Ag-ELISA. Each carcase was examined for the presence of Taenia hydatigena cysticerci and half carcase musculature together with the whole brain, head muscles, tongue, heart and diaphragm were sliced with fine cuts (< 0.5 cm) to reveal and enumerate T. solium cysticerci. Half carcase dissection can detect at least 84% of infected pigs. Prevalence and their 95% confidence intervals (CI) were calculated in Stata 12. Sensitivity, specificity, predictive values and likelihood ratios were determined. Results Twenty–nine pigs (8.3%, 95% CI: 5.6–11.7%) had viable T. solium cysticerci while 11 pigs had T. hydatigena cysticerci (3.1%, 95% CI: 1.6–5.5%). No co-infection was observed. Sixty-eight pigs (19.4%, 95% CI: 15.4–20%) tested positive on Ag-ELISA; of these, 24 had T. solium cysticerci and 7 had T. hydatigena cysticerci. Sensitivity and specificity were determined to be 82.7% and 86.3%, respectively. Positive and negative predictive values were 35.2% and 98.2%, respectively. Likelihood ratios for positive and negative Ag-ELISA test results were 6.0 and 0.2, respectively. There was a significant positive correlation between the titre of circulating antigens and intensity of T. solium cysticerci (r(348) = 0.63, P < 0.001). Conclusions The Ag-ELISA test characteristics reported in this study indicate that the test is more reliable in ruling out T. solium cysticercosis in pigs, than in confirming it. Hence, a negative result will almost certainly indicate that a pig has no infection, but a positive result should always be interpreted with caution. Estimates of T. solium prevalence based on Ag-ELISA results should, therefore, be adjusted for test performance characteristics and occurrence of T. hydatigena.


2015 ◽  
Vol 23 (3) ◽  
pp. 443-450 ◽  
Author(s):  
Jossuel Carvalho Melo Martins ◽  
Marcelo Maroco Cruzeiro ◽  
Leopoldo Antônio Pires

Objetivo. Relatar um caso de coinfecção por neurotoxoplasmose e neurocisticercose em mulher acometida pela AIDS. Método. Relato de caso prospectivo, descritivo e contemporâneo de paciente do sexo feminino, 36 anos, com quadro clínico compatível com síndrome de hipertensão intracraniana. Os exames complementares diagnostica­ram neurotoxoplasmose e a sorologia anti-HIV foi positiva. Evolui, após semanas, com crises convulsivas e exames subsidiários de imagem demonstrando neurocisticercose. Resultados. As infecções oportunis­tas relacionadas à AIDS são frequentes. A neurotoxoplasmose é cau­sada pelo protozoário Toxoplasma gondii e principal causa de lesão intracraniana expansiva em pacientes com AIDS. A neurocisticercose, provocada por cisticercos (Cysticercus cellulosae ou C. racemosus), formas larvárias da Taenia solium, é bastante prevalente em nosso meio. Conclusão. Após suspeição clínico-radiológica de neurotoxo­plasmose, torna-se imperativo realização de teste anti-HIV, devido sua elevada frequência neste grupo de pacientes. Já a comorbidade neurocisticercose e AIDS é achado excepcional, resultado mais prova­velmente de mera coincidência, sem qualquer vínculo predisponente entre si.


2013 ◽  
Vol 20 (8) ◽  
pp. 1143-1149 ◽  
Author(s):  
Weiwei Yan ◽  
Muhammad Hassan Saleem ◽  
Patrick McDonough ◽  
Sean P. McDonough ◽  
Thomas J. Divers ◽  
...  

ABSTRACTLeptospiraimmunoglobulin (Ig)-like (Lig) proteins are a novel family of surface-associated proteins in which the N-terminal 630 amino acids are conserved. In this study, we truncated the LigA conserved region into 7 fragments comprising the 1st to 3rd (LigACon1-3), 4th to 7.5th (LigACon4-7.5), 4th (LigACon4), 4.5th to 5.5th (LigACon4.5–5.5), 5.5th to 6.5th (LigACon5.5–6.5), 4th to 5th (LigACon4-5), and 6th to 7.5th (LigACon6-7.5) repeat domains. All 7 recombinant Lig proteins were screened using a slot-shaped dot blot assay for the diagnosis of equine leptospirosis. Our results showed that LigACon4-7.5 is the best candidate diagnostic antigen in a slot-shaped dot blot assay. LigACon4-7.5 was further evaluated as an indirect enzyme-linked immunosorbent assay (ELISA) antigen for the detection ofLeptospiraantibodies in equine sera. This assay was evaluated with equine sera (n= 60) that were microscopic agglutination test (MAT) negative and sera (n= 220) that were MAT positive to the 5 serovars that most commonly cause equine leptospirosis. The indirect ELISA results showed that at a single serum dilution of 1:250, the sensitivity and specificity of ELISA were 80.0% and 87.2%, respectively, compared to those of MAT. In conclusion, an indirect ELISA was developed utilizing a recombinant LigA fragment comprising the 4th to 7.5th repeat domain (LigACon4-7.5) as a diagnostic antigen for equine leptospirosis. This ELISA was found to be sensitive and specific, and it yielded results that concurred with those of the standard MAT.


2015 ◽  
Vol 24 (4) ◽  
pp. 471-474 ◽  
Author(s):  
Hanstter Hallison Alves Rezende ◽  
Juliana Boaventura Avelar ◽  
Heloísa Ribeiro Storchilo ◽  
Marina Clare Vinaud ◽  
Ana Maria de Castro

Abstract The accuracy of the parasitological techniques of Willis, Hoffman-Pons-Janer or Lutz (HPLJ), Sheather and Faust was evaluated in fecal samples from stray cats caught by the Zoonosis Control Center in Goiânia, Goiás, Brazil. These four techniques were applied separately to analyze 154 fecal samples, and their accuracy was analyzed based on an evaluation of their sensitivity, specificity, positive and negative predictive values, and Kappa index, resulting in the selection of the Willis technique as the nominal gold standard. Of the 154 samples, 115 (74.68%) tested positive for intestinal parasites. The analysis of the frequency of positivity indicated that the HPLJ technique detected 86.1% of the positive samples and was the closest to the gold standard. The analysis of the accuracy of the techniques was evaluated using the most prevalent parasites. The Sheather technique showed the highest accuracy in the detection of Ancylostomatidae, while the Sheather and HPLJ techniques showed similar accuracies in the detection of Cystoisospora spp. when compared to the gold standard. Lastly, the Faust technique showed the highest accuracy in the detection of Toxoplasma gondii when compared to the gold standard. This study underscores the importance of combining parasitological techniques in the diagnosis of intestinal parasites in cats.


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