scholarly journals Development and evaluation of an indirect ELISA for detection of Teladorsagia circumcincta infection in sheep

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Jalal Aliabadi ◽  
Ehsan Rakhshandehroo ◽  
Azadeh Yektaseresht
Keyword(s):  
Indirect Elisa ◽  
High Specificity ◽  
Negative Control ◽  
Infected Sheep ◽  
Diagnostic Methods ◽  
Elisa Method ◽  
Specific Product ◽  
Molecular Weights ◽  
Teladorsagia Circumcincta ◽  
Test Specificity

Abstract Background The gastrointestinal helminth, Teladorsagia circumcincta, is one of the major health risks and production-limiting diseases in small ruminant populations, particularly in temperate regions. With the increasing importance of disease management and recruited anthelmintic resistant types, accurate approaches are needed for the diagnosis of the infection in the host. Due to uncertain results using faecal examinations, the ELISA method was indicated for the detection of nematode antigenic materials. Despite some promising results, problems were described in terms of test specificity and cross-reactions. Therefore, this study aimed to evaluate the IgG response to worm somatic and excretory/secretory (ES) products using western blot analysis and an indirect ELISA for the detection of T. circumcincta infection in sheep. Results Based on the immuno-reactivity analysis, immunogenic fractions with molecular weights (MWs) of approximately 60, 75 and 100 kDa were detected in somatic content and two antigens of about 63 and 75 kDa in ES material. Accordingly, a specific product at 75 kDa had the strongest reaction and appeared as the most common antigenic protein. In ELISA, all the sera from the infected sheep revealed the OD rates above the calculated cut-off value with about two-fold greater average. Negative control samples were also specifically recognized with the mean OD rate of about 1/3 of the estimated cut-off value. The cross-reaction test, using rabbit anti-T. circumcincta IgG, did not show reactivity with the ES antigens of other prevalent nematodes including Haemonchus contortus, Protostrongylus rufescens and Marshallagia marshalli. In contrast, a strong positive reaction was observed with the somatic antigens of M. marshalli. Conclusions The results of this study indicated that the indirect ELISA method using the ES content enables distinguishing the T. circumcincta infected sheep with high specificity. Those antigenic ES peptides with 63 and particularly 75 kDa MWs should be further investigated due to the potential for serological diagnostic methods and immunoprotective targets in the host.

scholarly journals Development and Evaluation of an Indirect ELISA for Detection of Infection with Teladorsagia Circumcincta in Sheep

2021 ◽  
Author(s):  
Jalal Aliabadi ◽  
Ehsan Rakhshandehroo ◽  
Azadeh Yektaseresht
Keyword(s):  
Indirect Elisa ◽  
High Specificity ◽  
Negative Control ◽  
Infected Sheep ◽  
Diagnostic Methods ◽  
Elisa Method ◽  
Specific Product ◽  
Molecular Weights ◽  
Teladorsagia Circumcincta ◽  
The Cross

Abstract Background The gastrointestinal helminth, Teladorsagia circumcincta, is one of the major health risk and production-limiting disease in small ruminant populations, particularly in temperate regions. With the increasing importance of the disease management and recruited anthelminic resistant types, accurate approaches are needed for diagnosis of the infection in the host. Because of uncertain results using faecal examinations, the ELISA method was indicated for detection of the nematode antigenic materials. Despite some promising results, problems were described in terms of the test specificity and the cross-reactions. The aim of this study was therefore to evaluate worm somatic and excretory/secretory (ES) products using western blot analysis and an indirect ELISA for detecting T. circumcincta infection in sheep. Results Based on the immuno-reactivity analysis, immunogenic fractions with molecular weights of approximately 60, 75 and 100 kDa were detected in somatic content and two antigens of about 63 and 75 kDa in ES material. Accordingly, a specific product at 75 kDa had the strongest reaction and appeared as the most common antigenic protein. In ELISA, all sera from the infected sheep revealed the OD rates above the calculated cut-off value with about two-fold greater in average. Negative control samples were also specifically recognized with the mean OD rate of about 1/3 of the estimated cut-off value. The cross-reaction test, using rabbit hyperimmune sera, did not show reactivity with ES antigens of other prevalent nematodes include Haemonchus contortus, Protostrongylus rufescens and Marshallagia marshalli. In contrast, a strong positive reaction was found with the somatic antigens of M. marshalli. Conclusions The results obtained here indicates the indirect ELISA method using the ES content enables distinguishing the T. circumcincta infected sheep with high specificity. Those antigenic ES peptides with 63 and particularly 75 kDa molecular weights should be more investigated due to the potential for serological diagnostic methods and immunoprotective targets in the host.

scholarly journals Novel molecular transport medium used in combination with Xpert MTB/RIF ultra provides rapid detection of Mycobacterium bovis in African buffaloes

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Charlene Clarke ◽  
Katrin Smith ◽  
Samantha J. Goldswain ◽  
Christopher Helm ◽  
David V. Cooper ◽  
...  
Keyword(s):  
Mycobacterium Bovis ◽  
Human Health Risk ◽  
Molecular Transport ◽  
Sampling Technique ◽  
High Specificity ◽  
Negative Control ◽  
Post Mortem ◽  
Tissue Samples ◽  
Transport Medium ◽  
Tissue Homogenates

AbstractMycobacterium bovis is the causative agent of bovine tuberculosis (bTB) in wildlife. Confirmation of M. bovis infection relies on mycobacterial culture, which is time-consuming. Collection and transportation of infectious material also pose a human health risk. PrimeStore Molecular Transport Medium (MTM) has been shown to effectively inactivate infectious organisms, making it a safe method for handling infectious samples. This study investigated an in-field sampling technique for rapid, safe detection of M. bovis in buffalo tissues. Potentially infected tissues from bTB test-positive buffaloes were swabbed at post-mortem examination and stored in PrimeStore MTM at ambient temperature until Xpert MTB/RIF Ultra testing was performed. Additionally, tissue samples were frozen and transported before homogenisation for culture and Ultra testing. Oral swabs were collected from M. bovis-unexposed buffaloes as a negative control cohort. Mycobacterium tuberculosis complex (MTBC) DNA was detected by Ultra in 13/16 tissue swabs and 9/16 matched tissue homogenates from culture-confirmed M. bovis-positive buffalo tissues. MTBC DNA was not detected in swabs from M. bovis-unexposed animals, showing the potentially high specificity of Ultra with PrimeStore swabs. PrimeStore MTM sample processing, in combination with the Ultra assay, has the potential to provide a safe, rapid post-mortem screening test for M. bovis in buffaloes.

scholarly journals Crude antigen from Taenia crassiceps cysticercus used as heterologous antigen in ELISA and in EITB for neurocysticercosis diagnosis of patients from Paraná-Brazil

2008 ◽  
Vol 51 (6) ◽  
pp. 1127-1137 ◽  
Author(s):  
João Carlos Minozzo ◽  
Juliana de Moura ◽  
Sérgio Monteiro Almeida ◽  
Vanete Thomaz-Soccol
Keyword(s):  
Public Health ◽  
Neurological Disorders ◽  
Indirect Elisa ◽  
Taenia Solium ◽  
High Specificity ◽  
Heterologous Antigen ◽  
Public Health Systems ◽  
Crude Antigen ◽  
Specificity And Sensitivity ◽  
Cysticercus Cellulosae

Neurocysticercosis (NCC), the cerebral presence of Taenia solium metacestode (Cysticercus cellulosae), is responsible for neurological disorders worldwide. In order to validate an immunodiagnosis for public-health patients in the State of Parana-Brazil, crude antigen of Taenia crassicepsmetacestode (Cysticercus longicollis) was used as an alternative heterologous antigen to be used in ELISA and in electroimmunotransfer blotting (EITB) for active and inactive NCC diagnosis. Indirect ELISA was able to discriminate between active and inactive samples and presented high specificity and sensitivity. Any immunodominant band was able to distinguish the NCC stages, although the EITB showed 100% specificity. The immunological results proved to be an important auxiliary toll for NCC diagnosis, mainly for public-health systems in developing countries, where either the neuroimage techniques are not accessible or the resources are scarce.

scholarly journals Tagging of RPS9 as a tool for ribosome purification and identification of ribosome-associated proteins

2020 ◽  
pp. 57-57
Author(s):  
Bogdan Jovanovic ◽  
Lisa Schubert ◽  
Fabian Poetz ◽  
Georg Stoecklin
Keyword(s):  
Protein Synthesis ◽  
Ribosomal Proteins ◽  
Associated Factors ◽  
Ribosomal Subunit ◽  
High Specificity ◽  
Negative Control ◽  
Tev Protease ◽  
Protease Cleavage ◽  
Associated Proteins ◽  
And Function

Ribosomes, the catalytic machinery required for protein synthesis, are comprised of 4 ribosomal RNAs and about 80 ribosomal proteins in mammals. Ribosomes further interact with numerous associated factors that regulate their biogenesis and function. As mutations of ribosomal proteins and ribosome associated proteins cause many diseases, it is important to develop tools by which ribosomes can be purified efficiently and with high specificity. Here, we designed a method to purify ribosomes from human cell lines by C-terminally tagging human RPS9, a protein of the small ribosomal subunit. The tag consists of a flag peptide and a streptavidin-binding peptide (SBP) separated by the tobacco etch virus (TEV) protease cleavage site. We demonstrate that RPS9-Flag-TEV-SBP (FTS) is efficiently incorporated into the ribosome without interfering with regular protein synthesis. Using HeLa-GFP-G3BP1 cells stably expressing RPS9-FTS or, as a negative control, mCherry-FTS, we show that complete ribosomes as well as numerous ribosome-associated proteins are efficiently and specifically purified following pull-down of RPS9-FTS using streptavidin beads. This tool will be helpful for the characterization of human ribosome heterogeneity, post-translational modifications of ribosomal proteins, and changes in ribosome-associated factors after exposing human cells to different stimuli and conditions.

scholarly journals Performance evaluation of the Simtomax® CoronaCheck rapid diagnostic test

2020 ◽  
Author(s):  
P. J. Ducrest ◽  
A. Freymond ◽  
J.-M. Segura
Keyword(s):  
Diagnostic Test ◽  
Rapid Diagnostic Test ◽  
Diagnostic Performance ◽  
High Sensitivity ◽  
High Specificity ◽  
Negative Control ◽  
List Type ◽  
Plasma Samples ◽  
Rt Pcr ◽  
Two Samples

AbstractThe aim of this study was to evaluate the diagnostic performance of Simtomax® CoronaCheck, a serology rapid diagnostic test (RDT) for the detection of IgG and IgM against SARS-CoV-2. 48 plasma samples positive for SARS-CoV-2 based on RT-PCR and 98 negative control samples were studied. Diagnostic performance of the IgG/IgM RDT was assessed against RT-PCR and the electro-chemiluminescence immunoassay (ECLIA) Elecsys® Anti-SARS-CoV-2 total Ig. Overall, the RDT sensitivity was 92% (95% confidence interval [95%CI]: 79-97), specificity 97% (95% CI: 91-99%), PPV 94% (95% CI: 81-98) and the NPV 96% (95% CI: 89-99). When considering only samples collected ≥ 15 days post-symptoms (DPS), the sensitivity increased to 98% (95%CI: 86-100) and the specificity was 97% (95% CI: 91-99%). Two samples with 180 DPS were still positive for IgG. Globally, this IgG/IgM RDT displayed a high diagnostic accuracy for SARS-CoV-2 IgG/IgM detection in plasma samples in high COVID-19 prevalence settings. It could be effectively used, in absence of facilities for routine diagnostic serology, for samples with a DPS between 15 and 180 days.Highlights–The rapid diagnostic test Simtomax CoronaCheck displays a high sensitivity of 98% and a high specificity of 97% for SARS-CoV-2 IgG/IgM detection in plasma samples after 15 days post-symptoms.–The rapid diagnostic test Simtomax CoronaCheck can detect SARS-CoV-2 antibodies in plasma up to 180 days after symptom onset.–The rapid diagnostic test Simtomax CoronaCheck could be effectively used as an alternative to serological analysis using laboratory facilities.

scholarly journals Assessment of a recombinant protein from Leishmania infantum as a novel tool for Visceral Leishmaniasis (VL) diagnosis in VL/HIV co-infection cases

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0251861
Author(s):  
Rhaíssa E. M. Ramos ◽  
Wagner J. T. Santos ◽  
Franklin B. Magalhães ◽  
George T. N. Diniz ◽  
Carlos H. N. Costa ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Recombinant Protein ◽  
Indirect Elisa ◽  
Hiv Infections ◽  
Diagnostic Methods ◽  
Direct Agglutination Test ◽  
Serological Tests ◽  
Perfect Agreement ◽  
Significant Drop ◽  
Life Threatening

Visceral Leishmaniasis and HIV-AIDS coinfection (VL/HIV) is considered a life-threatening pathology when undiagnosed and untreated, due to the immunosuppression caused by both diseases. Serological tests largely used for the VL diagnosis include the direct agglutination test (DAT), ELISA and immunochromatographic (ICT) assays. For VL diagnosis in HIV infections, different studies have shown that the use of the DAT assay facilitates the VL diagnosis in co-infected patients, since the performance of the most widely used ELISA and ICT tests, based on the recombinant protein rK39, are much less efficient in HIV co-infections. In this scenario, alternative recombinant antigens may help the development of new serological diagnostic methods which may improve the VL diagnosis for the co-infection cases. This work aimed to evaluate the use of the recombinant Lci2 antigen, related to, but antigenically more diverse than rK39, for VL diagnosis in co-infected sera through ELISA assays. A direct comparison between recombinant Lci2 and rK39 was thus carried out. The two proteins were first tested using indirect ELISA with sera from VL afflicted individuals and healthy controls, with similar performances. They were then tested with two different sets of VL/HIV co-infected cases and a significant drop in performance, for one of these groups, was observed for rK39 (32% sensitivity), but not for Lci2 (98% sensitivity). In fact, an almost perfect agreement (Kappa: 0.93) between the Lci2 ELISA and DAT was observed for the coinfected VL/HIV patients. Lci2 then has the potential to be used as a new tool for the VL diagnosis of VL/HIV co-infections.

scholarly journals Comparison of Trichomoniasis Diagnosis by Microscopic Methods and Indirect ELISA Technique in a Sample of Iraqi Women

2020 ◽  
pp. 742-748
Author(s):  
Nada H. Bedair ◽  
Hayder Z. Ali
Keyword(s):  
Trichomonas Vaginalis ◽  
Vaginal Discharge ◽  
Indirect Elisa ◽  
Elisa Method ◽  
Transmitted Infection ◽  
Sexually Transmitted ◽  
Wet Mount ◽  
Urine Examination ◽  
Iraqi Women ◽  
Elisa Technique

Trichomonas vaginalis is an eukaryotic parasite that causes the most common non-viral sexually transmitted infection, trichomoniasis. This disease is responsible for many serious health problems such as preterm birth. More than half of the infected women do not develop symptoms, which makes it difficult to diagnose thedisease. In this study, a specific indirect ELISA method was developed to detect anti-Trichomonas vaginalis IgM and IgG immunoglobulins in the sera of infected females. The aim of this study was to investigate the sensitivity of a simple ELISA procedure in comparison to the classical urine examination and vaginal wet mount preparation for the diagnosis of T. vaginalis. The sensitivity of the indirect ELISA was compared with the classical vaginal discharge swab and urine microscopic examination, and the results showed sensitivities of 65.5% and 57.2%, respectively. Furthermore, the infection was measurable as acute or chronic with the refined test.

What are the best diagnostic tests for diagnosing bacterial arthritis of a native joint?

2021 ◽  
Vol 103-B (12) ◽  
pp. 1745-1753
Author(s):  
Alex B. Walinga ◽  
Tobias Stornebrink ◽  
David W. G. Langerhuizen ◽  
Peter A. A. Struijs ◽  
Gino M. M. J. Kerkhoffs ◽  
...  
Keyword(s):  
Synovial Fluid ◽  
Diagnostic Tests ◽  
High Specificity ◽  
Diagnostic Methods ◽  
Youden Index ◽  
Polymorphonuclear Cells ◽  
Gram Stain ◽  
Bacterial Arthritis ◽  
Diagnostic Modality ◽  
Serum Crp

Aims This study aimed to answer two questions: what are the best diagnostic methods for diagnosing bacterial arthritis of a native joint?; and what are the most commonly used definitions for bacterial arthritis of a native joint? Methods We performed a search of PubMed, Embase, and Cochrane libraries for relevant studies published between January 1980 and April 2020. Of 3,209 identified studies, we included 27 after full screening. Sensitivity, specificity, area under the curve, and Youden index of diagnostic tests were extracted from included studies. We grouped test characteristics per diagnostic modality. We extracted the definitions used to establish a definitive diagnosis of bacterial arthritis of a native joint per study. Results Overall, 28 unique diagnostic tests for diagnosing bacterial arthritis of a native joint were identified. The following five tests were deemed most useful: serum ESR (sensitivity: 34% to 100%, specificity: 23% to 93%), serum CRP (sensitivity: 58% to 100%, specificity: 0% to 96%), serum procalcitonin (sensitivity: 0% to 100%, specificity: 68% to 100%), the proportion of synovial polymorphonuclear cells (sensitivity: 42% to 100%, specificity: 54% to 94%), and the gram stain of synovial fluid (sensitivity: 27% to 81%, specificity: 99% to 100%). Conclusion Diagnostic methods with relatively high sensitivities, such as serum CRP, ESR, and synovial polymorphonuclear cells, are useful for screening. Diagnostic methods with a relatively high specificity, such as serum procalcitonin and synovial fluid gram stain, are useful for establishing a diagnosis of bacterial arthritis. This review helps to interpret the value of various diagnostic tests for diagnosing bacterial arthritis of a native joint in clinical practice. Cite this article: Bone Joint J 2021;103-B(12):1745–1753.

scholarly journals Immunohistochemical detection of soya protein – optimisation and verification of the method

2009 ◽  
Vol 27 (No. 1) ◽  
pp. 11-19 ◽  
Author(s):  
M. Pospiech ◽  
B. Tremlová ◽  
E. Renčová ◽  
Z. Randulová
Keyword(s):  
Indirect Elisa ◽  
Soya Protein ◽  
Immunohistochemical Method ◽  
Immunohistochemical Detection ◽  
Elisa Method ◽  
Background Staining ◽  
Soya Proteins ◽  
Abc Method ◽  
Soya Isolate ◽  
Avidin Biotin Complex

A functional immunohistochemical method for soya proteins detection was developed. The procedure is based on the avidin-biotin complex (ABC) method that attains sufficient sensitivity. The method was verified by the analysis of the model samples of different forms of soya additives containing various concentrations of soya isolate. The detection limit of soya present in the model samples was 0.5%. Different possibilities of the background staining were tested. The best results were obtained with the background staining according to Calleja. The results were confirmed by the accredited indirect ELISA method. The method allows the identification of various forms of soya proteins such as isolates, texturates, concentrates, and flour.

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