scholarly journals Performance of a 27 kDa Fasciola hepatica Antigen in the Diagnosis of Human Fascioliasis

2015 ◽  
Vol 7 (01) ◽  
pp. 017-020 ◽  
Author(s):  
Reza Shafiei ◽  
Bahador Sarkari ◽  
Seyed Mahmoud Sadjjadi
Keyword(s):  
Sensitivity And Specificity ◽  
Western Blotting ◽  
Indirect Elisa ◽  
Fasciola Hepatica ◽  
Serological Diagnosis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Healthy Controls ◽  
Elisa System ◽  
Immunodominant Antigen ◽  
Human Fascioliasis

ABSTRACT Background: Serological diagnosis, based on antigenic fractions of the parasite can be used for the early diagnosis of human fascioliasis. The current study aimed to evaluate the efficacy of a 27 kDa immunodominant antigen of Fasciola hepatica adult worms, in an indirect enzyme-linked immunosorbent assay (ELISA) system for serological diagnosis of human fascioliasis. Materials and Methods: The immunodiagnosis of human fascioliasis, using a 27 kDa immunodominant antigen, purified from F. hepatica somatic antigens (SAs), was evaluated by Western blotting and ELISA with sera samples of human fascioliasis patients, healthy controls and patients with other parasitic infections. Results: Using western blotting, from 12 sera of fascioliasis patients, 11 sera (91.6%) detected the 27 kDa subunit. None of 30 samples from healthy controls or 32 sera from nonfascioliasis patients reacted with the 27 kDa antigen. Accordingly, sensitivity and specificity of the system was found to be 91.6% and 100%, respectively. The 27 kDa antigen was purified from the SAs and was used in an indirect ELISA system. Of 15 sera of fascioliasis patients, all (100%) were found to be positive by ELISA whereas only 4 cases (6.25%) of nonfascioliasis patients or healthy controls were false-positive by this system. Accordingly, the sensitivity and specificity of the test were 100% and 93.6%, respectively. Conclusion: Findings of this study demonstrated that both Western blotting and the indirect ELISA, based on the 27 kDa subunit of F. hepatica SA, are reliable methods for serodiagnosis of human fascioliasis.

scholarly journals Combined Use of Indirect ELISA and Western Blotting with Recombinant Hepatocellular Carcinoma-Associated Antigen 59 Is a Potential Immunodiagnostic Tool for the Detection of Prepatent Haemonchus contortus Infection in Goat

Animals ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 548 ◽  
Author(s):  
Muhammad Ali-ul-Husnain Naqvi ◽  
Sana Zahra Naqvi ◽  
Muhammad Ali Memon ◽  
Kalibixiati Aimulajiang ◽  
Muhammad Haseeb ◽  
...  
Keyword(s):  
Western Blot ◽  
Western Blotting ◽  
Haemonchus Contortus ◽  
Indirect Elisa ◽  
Fasciola Hepatica ◽  
False Negative ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Combined Use

Haemonchus contortus is recognized as one of the important health problems in small ruminants, leading to reduced production and economic loss for farmers worldwide. Prepatent diagnosis of H. contortus infection is crucial to improve control strategies as this helminth may remove up to one-fifth of total erythrocytes and may cause anemia, edema, diarrhea, and ultimately death in young animals. In this study, one of the excretory and secretory products, rHc-HCA59, was purified and used as antigen to detect specific antibodies in H. contortus infected goats during prepatent stage of infection using indirect enzyme linked immunosorbent assay (ELISA) as screening test. All goats (n = 38) were housed indoor, experimentally infected with 8000 infective larvae (L3) of H. contortus, and serum samples were collected prior to infection and at 14th day of infection. Immunoblotting was performed to confirm the results of indirect ELISA, evaluate the cross reactivity against rHc-HCA59 in sera of most common co-infecting parasites and rectify the false negative samples. Furthermore, three different batches of rHc-HCA59 were produced to evaluate the repeatability of ELISA. No eggs were detected in feces of all goats collected at 7th and 14th day of infection but, H. contortus eggs were detected at 21 days post infection in the feces. Indirect ELISA performed in this study showed 87% sensitivity and 100% specificity. The western blot analysis confirmed immunoreactivity in serum samples which scored positive in indirect ELISA and recognized the samples as negative which had OD450 lower than negative cut-off value in indirect ELISA. Furthermore, all false negative sera (n = 5) that had OD450 value between positive and negative cut-off value in rHc-HCA59 based ELISA were clearly positive in western blot. Moreover, no cross-reactivity was detected in ELISA and western blotting against rHc-HCA59 in positive sera of Toxoplasma gondii, Fasciola hepatica, and Trichinella spiralis. The results of this study concluded that combined use of indirect ELISA and western blotting with rHc-HCA59 is a potential immunodiagnostic tool for the detection of H. contortus infection during prepatent period in goats.

scholarly journals Diagnosis of Visceral Leishmaniasis by Enzyme-Linked Immunosorbent Assay Using Urine Samples

2002 ◽  
Vol 9 (4) ◽  
pp. 789-794 ◽  
Author(s):  
Mohammad Zahidul Islam ◽  
Makoto Itoh ◽  
S. M. Shamsuzzaman ◽  
Rusella Mirza ◽  
Farzana Matin ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Sensitivity And Specificity ◽  
Immunosorbent Assay ◽  
Leishmania Donovani ◽  
Laboratory Diagnosis ◽  
Urine Samples ◽  
Enzyme Linked Immunosorbent Assay ◽  
Healthy Controls ◽  
Serum Samples ◽  
Crude Antigen

ABSTRACT A diagnostic method has been developed to detect anti-Leishmania donovani immunoglobulin G (IgG) in urine by enzyme-linked immunosorbent assay (ELISA). In measuring anti-L. donovani IgG, IgA, and IgM in urine, the method performed best in the detection of IgG. The sensitivity and specificity of the assay were determined with panels of urine samples from 62 visceral leishmaniasis (VL) patients, 59 healthy controls from areas of endemicity, 53 healthy controls from areas of nonendemicity, 59 malaria patients, 13 tuberculosis patients, 23 cutaneous leishmaniasis patients, and 7 patients with other diseases. Using L. donovani promastigote crude antigen, the test had 93.5% sensitivity (58 positives of 62 VL patient samples) and 89.3% specificity (191 negatives of 214 non-VL patient samples). The ELISA with acetone-treated L. donovani promastigote antigen raised the sensitivity and specificity to 95.0 and 95.3%, respectively. Western blot analysis revealed that most of the samples that cross-reacted with crude antigen in ELISA did not recognize any antigenic component of L. donovani crude antigen. We also checked 40 serum samples from the same group of VL patients for anti-L. donovani IgG and got 90.0% sensitivity with both crude and acetone-treated antigens. As collection of urine is much easier than collection of serum, the detection of anti-L. donovani IgG in urine with acetone-treated antigen will be useful in epidemiological studies. It could be an adjunct of laboratory diagnosis.

scholarly journals Leucine Aminopeptidase Is an Immunodominant Antigen of Fasciola hepatica Excretory and Secretory Products in Human Infections

2007 ◽  
Vol 15 (1) ◽  
pp. 95-100 ◽  
Author(s):  
A. Marcilla ◽  
J. E. De la Rubia ◽  
J. Sotillo ◽  
D. Bernal ◽  
C. Carmona ◽  
...  
Keyword(s):  
Phosphoenolpyruvate Carboxykinase ◽  
Leucine Aminopeptidase ◽  
Fasciola Hepatica ◽  
Enzyme Linked Immunosorbent Assay ◽  
Strong Reaction ◽  
Immunodominant Antigen ◽  
Proteomic Approach ◽  
Immunodominant Antigens ◽  
Secretory Products ◽  
Human Infections

ABSTRACT The liver fluke Fasciola hepatica parasitizes humans and ruminant livestock worldwide, and it is now being considered a reemerging zoonotic disease, especially in areas in which it is endemic, such as South America. This study investigates the immune response to excretory and secretory products produced by F. hepatica in a group of patients from the Peruvian Altiplano, where the disease is highly endemic. Using a proteomic approach and immunoblotting techniques, we have identified the enzymes leucine aminopeptidase (LAP) and phosphoenolpyruvate carboxykinase as immunodominant antigens recognized by sera from fasciolosis patients. An indirect enzyme-linked immunosorbent assay using recombinant LAP as the antigen was developed to check sera from individuals of this region. Our results demonstrate that LAP produces a specific and strong reaction, suggesting its potential use in the serologic diagnosis of F. hepatica infections in humans.

scholarly journals Increased miR-25 expression in serum of gastric cancer patients is correlated with CA19-9 and acts as a potential diagnostic biomarker

Open Medicine ◽  
2017 ◽  
Vol 12 (1) ◽  
pp. 266-270
Author(s):  
Qin Le ◽  
Niu Jianhua ◽  
Xi Yu ◽  
He Jiageng ◽  
Keyword(s):  
Gastric Cancer ◽  
Sensitivity And Specificity ◽  
Roc Curve ◽  
Diagnostic Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Healthy Controls ◽  
Diagnostic Biomarker ◽  
Relative Expression ◽  
Potential Biomarker ◽  
Diagnostic Sensitivity And Specificity

AbstractObjectiveTo evaluate miR-25 expression in serum of gastric cancer (GC) patients and whether it can be a potential biomarker for GC diagnosis.MethodsForty one pathology confirmed GC patients and 41 healthy controls were included in this study. 10 mL peripheral venous blood were collected from GC patients and healthy controls. miR-25 relative expression and CA19-9 level in serum of GC patients and healthy controls were measured by real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR) and enzyme linked immunosorbent assay (ELISA), respectively. The diagnostic sensitivity, specificity and receiver operating characteristic (ROC) curve of serum miR-25 and CA19-9 were calculated by STATA11.0 software.ResultsThe relative expression of miR-25 was 0.47±0.53 and 0.05±0.36 in serum of GC patients and healthy controls respectively with significant statistical difference (P<0.05). The serum level of CA19-9 for GC patients and healthy controls were 11.91±6.17 U/mL and 7.40±3.97 U/mL, indicating GC patients were much higher than those of healthy controls (P<0.05). The diagnostic sensitivity and specificity were 78.05% and 60.98% with the cut-off value of 0.32 for serum miR-25. Under this cut-off value, the area under the ROC curve was 0.75. For serum CA19-9, the diagnostic sensitivity and specificity were 70.73% and 56.10% with the cut-off value of 9.22 U/mL. Under this cut-off value, the area under the ROC curve was 0.73 with the 95% confidence interval of 0.62-0.84. Positive correlation was found between serum miR-25 relative expression and CA19-9 concentration of GC patients (r=0.75, P<0.05).ConclusionAccording to our present study, serum miR-25 was elevated in GC patients which may serve as a diagnostic biomarker for GC.

scholarly journals Comparative Performance of In-House and Commercially Available ELISA Kits for the Detection of Antibody Against Newcastle Disease Virus Vaccine

2013 ◽  
Vol 22 (1-2) ◽  
pp. 55-64
Author(s):  
MMI Chowdhury ◽  
MT Islam ◽  
A Aktar ◽  
MKJ Bhuiyan ◽  
MM Kamal ◽  
...  
Keyword(s):  
Newcastle Disease Virus ◽  
Disease Virus ◽  
Sensitivity And Specificity ◽  
Newcastle Disease ◽  
Indirect Elisa ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Dilution ◽  
Positive Correlation ◽  
Elisa Kit

An In-house Indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of serum antibody titre against Newcastle disease virus (NDV) and was compared its sensitivity and specificity with the commercially available NDV antibody detection ELISA kit (Biocheck®, USA). The reference NDV was purified by centrifugation, ultracentrifugation and by sucrose density gradient ultracentrifugation. This purified NDV was used for coating of 96-well flat bottomed microtitre plate and to raise hyperimmune sera (known) in Fayoumi chickens. In the standardization test, the antigen dilution of 10-6 and the serum dilution of 10-3 were considered to be optimum for the present ELISA system. The correlation regression analysis was performed to construct a standard curve equation where a good positive correlation was observed (r = 0.912, n = 8, P<0.01). The equation was used to convert corrected absorbance readings of the single working dilution (1 : 1000) directly into predicted ELISA antibody activity titres. In the sensitivity and specificity test, the serum dilution of 10-5 appeared to be the highest dilution which had the maximum lowest capacity to bind with the coated antigen of the present ELISA kit and only anti-NDV serum was found to bind with the coated antigen instead of serum of IBDV in the plate which revealed the high specificity of the developed In-house Indirect ELISA kit. In a comparative study with the 80 chicken sera samples, a significant positive correlation (r = 0.901, n = 80, P<0.01) was found between In-house Indirect ELISA and commercial ELISA kit (Biocheck®, USA).DOI: http://dx.doi.org/10.3329/pa.v22i1-2.16467 Progress. Agric. 22(1 & 2): 55 - 64, 2011

scholarly journals Immunodiagnosis of Human Fascioliasis by an Enzyme-Linked Immunosorbent Assay (ELISA) and a Micro-ELISA

2001 ◽  
Vol 8 (1) ◽  
pp. 174-177 ◽  
Author(s):  
Silvana Carnevale ◽  
Mónica I. Rodrı́guez ◽  
Graciela Santillán ◽  
Jorge H. Labbé ◽  
Marta G. Cabrera ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Fasciola Hepatica ◽  
Enzyme Linked Immunosorbent Assay ◽  
Screening Assay ◽  
Human Fascioliasis ◽  
Secretory Antigen ◽  
Excretory Secretory Antigen

ABSTRACT Enzyme-linked immunosorbent assay (ELISA) and micro-ELISA were evaluated for their ability to detect anti-Fasciola hepatica antibodies in humans by using excretory-secretory antigen. The sensitivity of each method was 100%, but the specificity was 100% for ELISA and 97% for micro-ELISA. The micro-ELISA could be used as a screening assay and ELISA could be used as a confirmatory method for the serodiagnosis of human fascioliasis.

scholarly journals Detection of Antibodies to BrucellaCytoplasmic Proteins in the Cerebrospinal Fluid of Patients with Neurobrucellosis

1999 ◽  
Vol 6 (5) ◽  
pp. 756-759 ◽  
Author(s):  
Pablo C. Baldi ◽  
George F. Araj ◽  
Graciela C. Racaro ◽  
Jorge C. Wallach ◽  
Carlos A. Fossati
Keyword(s):  
Cerebrospinal Fluid ◽  
Western Blotting ◽  
Immunosorbent Assay ◽  
Immunoglobulin G ◽  
Serological Diagnosis ◽  
Neurological Involvement ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Diagnosis ◽  
Serum Igg ◽  
Cytoplasmic Proteins

ABSTRACT The diagnosis of human neurobrucellosis usually relies on the detection of antibodies to Brucella lipopolysaccharide (LPS) in cerebrospinal fluid (CSF) by agglutination tests or enzyme-linked immunosorbent assay (ELISA). Here we describe the detection of immunoglobulin G (IgG) to cytoplasmic proteins (CP) ofBrucella spp. by ELISA and Western blotting in seven CSF samples from five patients with neurobrucellosis. While IgG to CP (titers of 200 to 12,800) and IgG to LPS (800 to 6,400) were found in the CSF of these patients, these antibodies were not detected in CSF samples from two patients who had systemic brucellosis without neurological involvement. The latter, however, had serum IgG and IgM to both LPS and CP. No reactivity to these antigens was found in CSF samples from 14 and 20 patients suffering from nonbrucellar meningitis and noninfectious diseases, respectively. These findings suggest that, in addition to its usefulness in the serological diagnosis of human systemic brucellosis, the ELISA with CP antigen can be used for the specific diagnosis of human neurobrucellosis.

scholarly journals Evaluation of an in-house indirect enzyme-linked immunosorbent assay of feline panleukopenia VP2 subunit antigen in comparison to hemagglutination inhibition assay to monitor tiger antibody levels by Bayesian approach

2020 ◽  
Author(s):  
Chanakan Areewong ◽  
Amarin Rittipornlertrak ◽  
Boondarika Nambooppha ◽  
Itsarapan Fhaikrue ◽  
Tawatchai Singhla ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Immunosorbent Assay ◽  
Hemagglutination Inhibition ◽  
Indirect Elisa ◽  
Enzyme Linked Immunosorbent Assay ◽  
Panthera Tigris ◽  
Serum Samples ◽  
Feline Panleukopenia Virus ◽  
Elisa Testing ◽  
Antibody Levels

Abstract Background: Feline panleukopenia virus (FPV) is an etiologic pathogen of feline panleukopenia that infects all members of Felidae including tigers (Panthera tigris). Vaccination against FPV among wild felid species has long been practiced in zoos worldwide. However, few studies have assessed tiger immune response post-vaccination due to the absence of a serological diagnostic tool. To address these limitations, this study aimed to develop an in-house indirect enzyme-linked immunosorbent assay (ELISA) for the monitoring of tiger antibody levels against the feline panleukopenia vaccine by employing the synthesized subunit capsid protein VP2. An in-house horseradish peroxidase (HRP) conjugated rabbit anti-tiger immunoglobulin G (IgG) polyclonal antibody (HRP-anti-tiger IgG) was produced in this study and employed in the assay. It was then compared to a commercial HRP-conjugated goat anti-cat IgG (HRP-anti-cat IgG). Sensitivity and specificity were evaluated using the Bayesian model with conditional dependence being assumed between both HRP-conjugated antibody-based ELISAs and hemagglutination-inhibition (HI) tests. Results: The posterior estimates for sensitivity and specificity of two indirect ELISA HRP-conjugated antibodies were higher than those of the HI test. The sensitivity and specificity of indirect ELISA for HRP-anti-tiger IgG and HRP-anti-cat IgG were 86.5%, 57.2% and 86.7%, 64.6%, respectively, while the results of the HI test were 79.1 and 54.1%. In applications, 89.6% (198/221) of tiger serum samples were determined to be seropositive by indirect ELISA testing. Conclusion: The results support evidence that an in-house indirect ELISA developed in this study could be used as a serological tool for the effective detection of tiger antibodies.

scholarly journals Evaluation of a Rapid Device for Serological Diagnosis of Leishmania infantum Infection in Dogs as an Alternative to Immunofluorescence Assay and Western Blotting

2013 ◽  
Vol 20 (5) ◽  
pp. 657-659 ◽  
Author(s):  
E. Ferroglio ◽  
S. Zanet ◽  
W. Mignone ◽  
M. Poggi ◽  
A. Trisciuoglio ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Western Blotting ◽  
Indirect Immunofluorescence ◽  
Leishmania Infantum ◽  
Immunofluorescence Assay ◽  
Serological Diagnosis ◽  
Immunochromatographic Test ◽  
Serum Samples ◽  
Content Type ◽  
Test Speed

ABSTRACTIn this study, we compared a rapid immunochromatographic test (Speed Leish K; BVT Groupe Virbac, La Seyne sur Mer, France) with an indirect immunofluorescence assay (IFAT) and Western blotting (WB) for the detection ofLeishmania infantumantibodies in dogs. A total of 250 serum samples were collected from 125L. infantum-positive and 125L. infantum-negative dogs. Among the positive samples, 81 were strongly positive at low IFAT dilutions, while 44 were low-reactivity sera (IFAT titers, 1:40 to 1:80). The sensitivity and specificity of the Speed Leish K were 96.3% and 100%, respectively, compared with those of the IFAT. When IFAT low-reactivity sera (titers, 1:40 or 1:80) were tested with the Speed Leish K, using WB results as a reference, the sensitivities were 93.75% for sera with a 1:80 titer and 73.33% for sera with a 1:40 titer, and the specificity was 100%. The Speed Leish K is easy to use and performs well, so it can be considered a quick and reliable tool for the diagnosis ofL. infantuminfection in dogs.

scholarly journals Development and Potential Application of Ras Domain Containing Protein from Haemonchus contortus for Diagnosis of Goat Infection

Animals ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 138 ◽  
Author(s):  
Kalibixiati Aimulajiang ◽  
Man Cao ◽  
Shuyi Liao ◽  
Muhammad Ali-ul-Husnain Naqvi ◽  
Xiaowei Tian ◽  
...  
Keyword(s):  
Haemonchus Contortus ◽  
Indirect Elisa ◽  
Fasciola Hepatica ◽  
Trichinella Spiralis ◽  
Clinical Signs ◽  
Small Ruminants ◽  
Gastrointestinal Nematode ◽  
Enzyme Linked Immunosorbent Assay ◽  
Fecal Examination ◽  
Diagnostic Potential

Haemonchus contortus is an important gastrointestinal nematode of small ruminants that causes significant mortality in goats worldwide. Diagnosis of this infection mainly depends on the evaluation of clinical signs and fecal examination. However, limitations often occur in early or mild infections. For this purpose, serological diagnosis seems to be more accurate and reliable. Ras domain-containing protein (Ras) is one of H. contortus’s excretory and secretory products (ESPs) that can be isolated from different larval stages of the nematode. In this study, the recombinant H. contortus Ras domain-containing protein (rHcRas) was expressed and purified and its diagnostic potential was evaluated. Reactions between rHcRas and goat sera were tested using Western blotting (WB). The results showed that rHcRas could be recognized by sera as early as 14 days post infection (DPI), and antibodies against rHcRas in infected goats could be maintained for over 89 days. No reaction was found between rHcRas and antibodies against Trichinella spiralis, Fasciola hepatica, or Toxoplasma gondii. An indirect enzyme-linked immunosorbent assay (ELISA) was produced based on rHcRas. The optimal coating antigen (157 ng of rHcRas/well) and serum dilutions (1:50) were determined via checkerboard titration. Indirect ELISA based on rHcRas showed 87.5% sensitivity and 90.6% specificity. The cut-off values for this experiment were determined to be 0.324 (positive) and 0.273 (negative), respectively, and the variation coefficient (CV) was less than 15%. The results of the indirect ELISA in-field examination showed that 17.6% (9/51) of the goats were infected with H. contortus, higher than the fecal examination results (15.7%, 8/51). When compared the results of the indirect ELISA and necropsy testing, 98.0% (50/51) consistency was found. These results indicated that rHcRas was a potential antigen for the diagnosis of H. contortus infection in goats.

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