scholarly journals Development of recombinant nucleocapsid protein-based indirect enzyme-linked immunosorbent assay for sero-survey of porcine reproductive and respiratory syndrome

2020 ◽  
Vol 13 (12) ◽  
pp. 2587-2595
Author(s):  
S. Phani Kashyap ◽  
Jagadish Hiremath ◽  
S. Vinutha ◽  
Sharanagouda S. Patil ◽  
Kuralayanapalya P. Suresh ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Disease Surveillance ◽  
Disease Outbreaks ◽  
Screening Tests ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
North East India ◽  
North East ◽  
Relative Specificity

Background and Aim: Porcine reproductive and respiratory syndrome (PRRS) is a disease endemic in many countries and is of economic importance. India was free from PRRS until the first outbreak was reported from a North-East Indian state in 2013. Since then, disease outbreaks have been reported from North-East India and the pilot study conducted earlier showed that it is gradually spreading to the rest of India. Considering there are no locally developed population screening tests available for PRRS and imported diagnostic/screening tests are expensive, the present study was aimed at developing recombinant nucleocapsid (rN) protein-based indirect enzyme-linked immunosorbent assay (iELISA). Materials and Methods: The rN protein of PRRS virus (PRRSV) was produced following standard cloning, expression, and purification procedures. Using this antigen, iELISA was optimized for the detection of serum antibodies to PRRSV. The sensitivity and specificity of the test were assessed by comparing it with a commercial PRRSV antibody detection kit. Results: A total of 745 serum samples from ten different states of India were screened using the developed iELISA. The iELISA had a relative specificity of 76.18% and sensitivity of 82.61% compared to the commercial ELISA (Priocheck PRRSV ELISA kit, Thermo Fisher Scientific, USA). Conclusion: The iELISA, which deployed rN protein from Indian PRRSV, was found to be suitable in the serological survey and may be a useful tool in future disease surveillance programs.

A study of the correlation between Coxiella burnetii seropositivity and abortions in sheep in Eastern and Southeastern Turkey

2016 ◽  
Author(s):  
Ayse Kilic ◽  
Hakan Kalender
Keyword(s):  
Immunosorbent Assay ◽  
Coxiella Burnetii ◽  
Q Fever ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Eastern Anatolia ◽  
Obligate Intracellular Bacterium ◽  
Serum Samples ◽  
Obligate Intracellular ◽  
Elisa Kit

Q fever is a zoonotic disease that occurs worldwide and is caused by the obligate intracellular bacterium Coxiella burnetii. Infected animals are usually asymptomatic, but infection can cause abortion and stillbirth in ruminants. The main purpose of this study was to evaluate prevalance of Coxiella burnetii infection in aborted and nonaborted sheep serum samples in Eastern Anatolia region by using enzyme-linked immunosorbent assay (ELISA). The determine of prevalance in sheep flocks from four provinces (Elazig, Malatya, Tunceli, Bitlis) and tested for anti-C.burnetii antibody detection, by means of Chekit Q fever Elisa kit. 350 serum samples obtained from flocks belonging aborted sheep showed that a total of 56 (16%) were detected seropositivity, whereas 171 serum samples obtained from nonaborted sheep flocks in 13 of the 171 (7.60%) for C.burnetii in seropositivity were observed. Coxiellosis should be considered an important cause of sheep with abortion history and nonaborted in Elazig and neighboring provinces.

scholarly journals Development of a Competitive Enzyme-Linked Immunosorbent Assay for Detection of Antibodies against the 3B Protein of Foot-and-Mouth Disease Virus

2015 ◽  
Vol 22 (4) ◽  
pp. 389-397 ◽  
Author(s):  
Ming Yang ◽  
Satya Parida ◽  
Tim Salo ◽  
Kate Hole ◽  
Lauro Velazquez-Salinas ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
Diagnostic Sensitivity ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Nonstructural Proteins ◽  
Serum Samples ◽  
Mouth Disease Virus ◽  
Foot And Mouth

ABSTRACTFoot-and-mouth disease (FMD) is one of the most highly contagious and economically devastating diseases, and it severely constrains the international trade of animals. Vaccination against FMD is a key element in the control of FMD. However, vaccination of susceptible animals raises critical issues, such as the differentiation of infected animals from vaccinated animals. The current study developed a reliable and rapid test to detect antibodies against the conserved, nonstructural proteins (NSPs) of the FMD virus (FMDV) to distinguish infected animals from vaccinated animals. A monoclonal antibody (MAb) against the FMDV NSP 3B was produced. A competitive enzyme-linked immunosorbent assay (cELISA) for FMDV/NSP antibody detection was developed using a recombinant 3ABC protein as the antigen and the 3B-specific MAb. Sera collected from naive, FMDV experimentally infected, vaccinated carrier, and noncarrier animals were tested using the 3B cELISA. The diagnostic specificity was 99.4% for naive animals (cattle, pigs, and sheep) and 99.7% for vaccinated noncarrier animals. The diagnostic sensitivity was 100% for experimentally inoculated animals and 64% for vaccinated carrier animals. The performance of this 3B cELISA was compared to that of four commercial ELISA kits using a panel of serum samples established by the World Reference Laboratory for FMD at The Pirbright Institute, Pirbright, United Kingdom. The diagnostic sensitivity of the 3B cELISA for the panel of FMDV/NSP-positive bovine serum samples was 94%, which was comparable to or better than that of the commercially available NSP antibody detection kits. This 3B cELISA is a simple, reliable test to detect antibodies against FMDV nonstructural proteins.

scholarly journals Rapid Competitive Enzyme-Linked Immunosorbent Assay for Detection of Antibodies to Peste des Petits Ruminants Virus

2005 ◽  
Vol 12 (4) ◽  
pp. 542-547 ◽  
Author(s):  
Kang-Seuk Choi ◽  
Jin-Ju Nah ◽  
Young-Joon Ko ◽  
Shien-Young Kang ◽  
Nam-In Jo
Keyword(s):  
Immunosorbent Assay ◽  
Small Ruminants ◽  
Viral Disease ◽  
Turnaround Time ◽  
Enzyme Linked Immunosorbent Assay ◽  
Peste Des Petits Ruminants ◽  
Rinderpest Virus ◽  
Serum Samples ◽  
Specificity And Sensitivity ◽  
Relative Specificity

ABSTRACT Peste des petits ruminants (PPR) is a contagious viral disease of small ruminants that is of economic importance in Africa, the Middle East, and Asia. We developed a rapid competitive enzyme-linked immunosorbent assay (rapid c-ELISA) for the diagnosis and surveillance of PPR. This assay detects PPR virus (PPRV) antibodies in serum samples by quantifying the amount of monoclonal antibody (MAb) P-3H12 after 30 min of incubation of a serum-MAb conjugate mixture on plates coated with a PPRV recombinant nucleocapsid protein (rPPRV-N). We tested 249 PPRV-positive serum samples and 733 PPRV-negative serum samples from field ruminants. The threshold of percent inhibition (PI) was determined to be <50 on the basis of the mean PI plus 3 standard deviations for sera from PPRV-negative ruminants. The relative specificity and sensitivity of the rapid c-ELISA were 98.5% (722 of 733 serum samples) and 93.4% (234 of 249 serum samples), respectively. The rapid c-ELISA sensitively detected PPRV antibodies in hyperimmune sera (virus neutralization test [VNT] titer, >512), even at dilutions ≥512 in normal goat serum, and as early as 6 to 13 days postinfection from 12 goats, each of which was infected with one of the four PPRV lineages. Hyperimmune sera from animals experimentally vaccinated with rinderpest virus gave positive results by the rapid c-ELISA when the rinderpest virus VNT titers were >512, although the rapid c-ELISA titers were very low (2 to 16). However, the rapid c-ELISA was negative when the rinderpest virus VNT titer was ≤128. The rapid c-ELISA developed in the present work provides a short turnaround time and could be a useful tool for the diagnosis of PPR and screening for PPRV in the field.

scholarly journals Serological Evaluation of Thin-Layer Immunoassay–Enzyme-Linked Immunosorbent Assay for Antibody Detection in Human Trichinellosis

2000 ◽  
Vol 7 (5) ◽  
pp. 810-812 ◽  
Author(s):  
Alberto Gómez-Priego ◽  
Lidia Crecencio-Rosales ◽  
Jorge-Luis de-la-Rosa
Keyword(s):  
Thin Layer ◽  
Immunosorbent Assay ◽  
Trichinella Spiralis ◽  
Field Studies ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Laboratory Equipment ◽  
Specialized Equipment ◽  
Human Trichinellosis

ABSTRACT A new immunoenzymatic test, named the thin-layer immunoassay–enzyme-linked immunosorbent assay (TIA-ELISA), was evaluated for antibody detection in human trichinellosis using excretion and secretion products prepared from Trichinella spiralis muscle larvae. Serum samples from people with positive muscle biopsies or symptoms compatible with the disease (n = 8 or 26, respectively), all reactive in enzyme-linked immunoelectrotransfer blot assay (EITB), as well as 67 serum samples from healthy, EITB-negative people, were tested in an ELISA and TIA-ELISA. TIA-ELISA was performed in polystyrene plastic petri dishes by adding dots of 10 μl each of antigen (7 μg/ml) followed by adding diluted serum and the conjugate. Finally, the substrate mixed with agar was added to develop the reaction. Enzymatic by-products were easily detected by the naked eye as defined dots. Sensitivity and specificity were 76 and 94% for ELISA, and both parameters were 91% for TIA-ELISA. The kappa correlation indices for both tests in relation to EITB were 0.73 and 0.80, respectively. The TIA-ELISA can be carried out with common laboratory equipment in 3 h and uses lower quantities of antigen than EITB and ELISA. Since TIA-ELISA is easy to perform, cheap, sensitive, and specific, the test could be an acceptable alternative to use in clinical laboratories lacking specialized equipment needed for ELISA and EITB and in field studies for antibody detection in human trichinellosis.

Development and application of an indirect enzyme-linked immunosorbent assay using recombinant S1 for serological testing of porcine epidemic diarrhea virus

2019 ◽  
Vol 65 (5) ◽  
pp. 343-352
Author(s):  
Ying Shan ◽  
Yajie Liu ◽  
Ziqi Liu ◽  
Guowei Li ◽  
Cong Chen ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Porcine Epidemic Diarrhea Virus ◽  
Fluorescent Antibody ◽  
Area Under The Curve ◽  
Antibody Detection ◽  
Economic Losses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Diarrhea Virus ◽  
Porcine Epidemic Diarrhea

Porcine epidemic diarrhea virus (PEDV) causes severe infectious diseases in all ages of swine and leads to serious economic losses. Serologic tests are widely accepted and used to detect anti-PEDV antibodies that could indicate PEDV infection or vaccination. In this study, PEDV recombinant S1 protein (rS1) was expressed with the Bac-to-Bac system and purified by nickel-affinity chromatography. An indirect enzyme-linked immunosorbent assay based on rS1 (rS1-ELISA) was then developed and optimized by checkerboard assays with serial dilutions of antigen and serum. Serum samples from 453 domestic pigs and 42 vaccinated pigs were analyzed by the indirect fluorescent antibody (IFA) test and rS1-ELISA. Taking IFA as a gold standard, rS1-ELISA produced a high sensitivity (90.7%) and specificity (94.6%) by a receiver operating characteristic (ROC) curve. In addition, ROC analysis also revealed that rS1-ELISA was consistent with IFA (area under the curve 0.9583 ± 0.0082). This rS1-ELISA was then applied to antibody detection in inactivated PEDV vaccinated pigs. The antibody could be detected 2–4 weeks after the first inoculation. These results indicated that the rS1-ELISA established in this study provides a promising and reliable tool for serologic detection of anti-PEDV IgG antibodies in infected or vaccinated pigs.

scholarly journals Fluorescence polarization assay, competitive enzyme-linked immunosorbent assay (ELISA-C) and indirect ELISA for the diagnosis of brucellosis in buffaloes (Bubalus bubalis)

2012 ◽  
Vol 42 (9) ◽  
pp. 1621-1626 ◽  
Author(s):  
Lília Márcia Silva Paulin ◽  
Luis Ernesto Samartino ◽  
Sandra Beatriz Conde ◽  
Igor Stefan Poppovic Federsoni ◽  
Fernando Ferreira ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Fluorescence Polarization ◽  
Relative Sensitivity ◽  
Bubalus Bubalis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Serological Tests ◽  
Relative Specificity ◽  
Fluorescence Polarization Assay ◽  
Kappa Agreement

The objective of the present study was to compare the performance of three serological tests for diagnosis of Brucella abortus infections in buffaloes (Bubalus bubalis). Serum samples collected from 696 adult females were submitted to the competitive enzyme-linked immunosorbent assay (ELISA-C), (I-ELISA), fluorescence polarization test (FPA), 2-mercaptoethanol test (2-ME) and complement fixation test (CFT). The gold standard was the combination of CFT and 2-ME, considering as positive the reactors in both CFT and 2-ME, and as negative those non-reactors. ROC analyses were done for C-ELISA, I-ELISA and FPA and the Kappa agreement index were also calculated. The best combinations of relative sensitivity (SEr) and relative specificity (SPr) and Kappa were given by C-ELISA (96.9%, 99.1%, and 0.932, respectively) and FPA (92.2%, 97.6 and 0.836, respectively). The C-ELISA and FPA were the most promising confirmatory tests for the serological diagnosis of brucellosis in buffaloes, and for these tests, cut-off values for buffaloes may be the same as those used for bovines.

scholarly journals Development of an Enzyme-Linked Immunosorbent Assay To Detect Antibodies Targeting Recombinant Envelope Protein 2 of Mayaro Virus

2019 ◽  
Vol 57 (5) ◽  
Author(s):  
Marcílio Jorge Fumagalli ◽  
William Marciel de Souza ◽  
Marília Farignoli Romeiro ◽  
Michell Charles de Souza Costa ◽  
Renata Dezengrini Slhessarenko ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Envelope Protein ◽  
Cross Reactivity ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
High Avidity ◽  
Serum Samples ◽  
Igg Antibody ◽  
Mayaro Virus ◽  
Recombinant Envelope Protein

ABSTRACT Mayaro virus (MAYV) is a neglected arthropod-borne virus (arbovirus) antigenically clustered into the Semliki Forest complex group of Alphavirus genus (Togaviridae family), maintained in an unclear zoonotic cycle involving mosquitoes from Haemagogus genus as the main vector. The genome is composed of a positive single-stranded RNA of 11.5 kb in length, which contains two genes that encode four nonstructural (nsP1 to nsP4) and five structural (C, E3, E2, 6K, and E1) proteins. In the present study, we have developed an enzyme-linked immunosorbent assay (ELISA) using as antigen the recombinant envelope protein 2 of MAYV produced in an Escherichia coli system (rE2-MAYV ELISAs). A panel of 68 human serum samples from suspected arboviral cases was analyzed and titrated for anti-MAYV IgM and IgG antibody detection. The rE2-MAYV ELISA detected 33.8% (23/68) IgG-positive samples, demonstrating 100% sensitivity and 78.95% specificity compared to the MAYV-specific 50% plaque reduction neutralization assay. In addition, the positive MAYV-neutralizing samples showed high titers of detection by rE2-MAYV ELISA, suggesting a highly sensitive test. The rE2-MAYV ELISA also detected 42.5% (29/68) IgM-positive samples, of which 13.8% (4/29) presented high-avidity interactions with rE2-MAYV. Cross-reactivity was observed with Chikungunya virus (CHIKV)-specific murine antibody sample but not with CHIKV-specific human and other Alphavirus murine antibodies. In short, we have developed a rapid, simple, specific, and sensitive MAYV rE2-ELISA, and our preliminary results show its potential applicability to diagnosis of MAYV infections.

DETECTION OF ANTI-RETICULOENDOTHELIOSIS ANTIBODY BY ENZYME-LINKED IMMUNOSORBENT ASSAY USING ENVELOPE PROTEIN EXPRESSED IN BACULOVIRUS

2016 ◽  
Vol 42 (03) ◽  
pp. 165-170
Author(s):  
Hui-Wen Chen ◽  
Ting-Hsiang Hsiao ◽  
Ching-Ho Wang
Keyword(s):  
Immunosorbent Assay ◽  
Envelope Protein ◽  
Insect Cells ◽  
Expression System ◽  
Baculovirus Expression System ◽  
Relative Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Baculovirus Expression ◽  
Relative Specificity

Reticuloendotheliosis virus (REV) infects various animals including chickens, ducks, geese, pheasants, peafowl and other birds. REV causes tumor, runting disease syndrome and immunosuppression in infected birds. The purpose of this study was to develop a rapid, reliable and convenient method to detect anti-REV antibody. The REV envelope protein was expressed by baculovirus expression system and evaluated for the potential as an antigen in an enzyme linked-immunosorbent assay (ELISA). The env gene from a REV strain goose/3410/06 was cloned into bacmid vector and expressed in Sf9 insect cells. The expected recombinant envelope protein expressed in infected insect cells was demonstrated in western blot with size of 62 kDa, and purified for using as antigen in an ELISA. A total of 182 chicken serum samples were used to evaluate the suitability of the ELISA based on neutralization test as the gold standard. The results showed that the relative sensitivity and relative specificity of this ELISA were 88.5% and 97.7%, respectively. The present ELISA using baculovirus expression envelope protein can be used to detect anti-REV antibody in the field.

scholarly journals Performance of a Johne's Disease Enzyme-Linked Immunosorbent Assay Adapted for Milk Samples from Goats

2005 ◽  
Vol 17 (4) ◽  
pp. 350-354 ◽  
Author(s):  
Miguel Salgado ◽  
Elizabeth J. B. Manning ◽  
Michael T. Collins
Keyword(s):  
Immunosorbent Assay ◽  
Low Cost ◽  
Goat Milk ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Prior History ◽  
Fecal Culture ◽  
Serum Samples ◽  
Milk Samples ◽  
Testing Cost

Antibody detection–based tests for paratuberculosis offer speed and economy, 2 diagnostic test attributes important to animal industries with narrow profit margins. Application of such tests to individual milk samples instead of serum samples can further improve testing efficiency and decrease testing cost. Accuracy of a commercial bovine paratuberculosis enzyme-linked immunosorbent assay (ELISA) adapted for use on goat serum and milk samples was determined. Fecal, blood, and milk samples were collected from 159 goats belonging to 2 Wisconsin goat herds with a prior history of paratuberculosis and 1 herd of 50 goats from a paratuberculosis-free Wisconsin herd. Fecal samples were cultured using the modified BACTEC 12B media. Sera were tested according to the manufacturer's instructions for bovine samples. Milk samples were centrifuged and mixed with the ELISA kit's Mycobacterium phlei–containing diluent at a ratio of 1:2. Using fecal culture as the “gold standard,” the sensitivity of the ELISA on goat serum was 64% and the sensitivity of the ELISA on goat milk was 48%. The milk ELISA had higher agreement with fecal culture results (kappa = 0.525) than the serum ELISA (kappa = 0.425). ELISA specificity was 100% on both serum and milk. Regression analysis also showed good correlation between serum and milk S/P values ( r2 = 0.67). Although less sensitive, the ELISA on goat milk samples appears to offer a useful, low-cost alternative for detection of goats with paratuberculosis that have progressed to the stage of shedding M. paratuberculosis in their feces.

scholarly journals Prevalence and clinical correlation of hepatitis E virus antibody in the patients’ serum samples from a tertiary care hospital in Thailand during 2015–2018

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Atiporn Boonyai ◽  
Anchalee Thongput ◽  
Thidarat Sisaeng ◽  
Parisut Phumchan ◽  
Navin Horthongkham ◽  
...  
Keyword(s):  
Pregnant Women ◽  
Tertiary Care ◽  
Laboratory Data ◽  
Organ Transplant ◽  
Hospital Setting ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Care Hospital ◽  
Serum Samples ◽  
Igm Antibodies

Abstract Background Prevalence and incidence of hepatitis caused by HEV infection are usually higher in developing countries. This study demonstrated the HEV seroprevalence and incidence of HEV infection in patients with clinical hepatitis in a tertiary hospital in Thailand. Methods A laboratory-based cross-sectional study was conducted using 1106 serum samples from patients suspected of HEV infection sent to the Serology laboratory, Siriraj Hospital, for detecting HEV antibodies during 2015–2018. Prevalence of anti-HEV IgG and IgM antibodies in general patients, including organ transplant recipients and pregnant women in a hospital setting, were determined using indirect enzyme-linked immunosorbent assay (ELISA) kits. Comparison of laboratory data between groups with different HEV serological statuses was performed. Results HEV IgG antibodies were detected in 40.82% of 904 serum samples, while HEV IgM antibodies were detected in 11.75% of 1081 serum samples. Similar IgG and IgM antibody detection rates were found in pregnant women. Interestingly, anti-HEV IgM antibodies were detected in 38.5% of patients who underwent organ transplantation. Patients who tested positive for anti-HEV IgM antibodies had higher alanine aminotransferase levels than those who had not. In contrast, patients who tested positive for anti-HEV IgG had more elevated levels of total bilirubin than those who tested negative. Conclusions HEV seroprevalence and incidence in patients with clinical hepatitis were relatively high in the Thai population, including the pregnancy and organ transplant subgroups. The results potentially benefit the clinicians in decision-making to investigate HEV antibodies and facilitating proper management for patients.

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