Evaluation of a Novel Enzyme-Linked Immunosorbent Assay for Detection of Antibodies against Salmonella, Employing a Stable Coating of Lipopolysaccharide-Derived Antigens Covalently Attached to Polystyrene Microwells

Polysaccharides derived from Salmonella typhimurium lipopolysaccharide (LPS) representing the O-antigen factors 1, 4, 5, and 12 and the O-antigen factors 6 and 7 from Salmonella choleraesuis LPS were derivatized with the photoreactive compound anthraquinone and subsequently covalently coupled to microtiter polystyrene plates by ultraviolet irradiation. Both polysaccharide antigens could be coupled simultaneously to the same microtiter plate. The coated surface was used in indirect ELISA for the determination of serum antibodies from pigs infected with bacteria of the two Salmonella groups and from uninfected pigs. This ELISA proved itself by having a good long-term durability and a high degree of reproducibility, including low day-to-day variations and low interplate variations. Furthermore, the ELISA showed good specificity and sensitivity when data were compared with the optical density levels of a panel of pig sera as determined by a conventional ELISA on the basis of passive coating of the two Salmonella LPS antigens (the mix-ELISA). The covalent anthraquinone mix-ELISA shows promise as a stable and durable alternative to the existing conventional ELISA for serological surveillance of Salmonella infections in pigs.

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Development and Evaluation of an Immunochromatographic Strip for Detection of Streptococcus Suis Type 2 Antibody

Journal of Veterinary Diagnostic Investigation ◽

10.1177/104063870701900403 ◽

2007 ◽

Vol 19 (4) ◽

pp. 355-361 ◽

Cited By ~ 15

Author(s):

Junxing Yang ◽

Meilin Jin ◽

Jianfeng Chen ◽

Ying Yang ◽

Pei Zheng ◽

...

Keyword(s):

Capsular Polysaccharide ◽

Protein A ◽

Streptococcus Suis ◽

Enzyme Linked Immunosorbent Assay ◽

Staphylococcal Protein A ◽

Immunochromatographic Strip ◽

Specificity And Sensitivity ◽

Antibody Titers ◽

Serological Surveillance ◽

And Control

In this study, an immunochromatographic strip (ICS) was developed for the detection of antibody against Streptococcus suis serotype 2 (SS2). Colloidal gold particles labeled with staphylococcal protein A (SPA), which can bind to the FC fragment of mammalian immunoglobulin, were used as the detector reagent. The capsular polysaccharide (CPS) of SS2 and affinity-purified IgG from a healthy naive pig were immobilized on test and control regions of a nitrocellulose membrane, respectively. The ICS was used to 1) detect anti-CPS antibody in 14 sera taken from 4 SS2-infected pigs, 24 sera from pigs hyperimmunized with SS2, and 68 sera from pigs inoculated or infected with bacteria other than SS2; 2) determine anti-CPS antibody titers of 20 positive sera for comparison with enzyme-linked immunosorbent assay (ELISA); and 3) detect anti-CPS antibody in 226 clinical sera taken from diseased pigs also for comparison with ELISA. An ELISA used as a reference test determined the specificity and sensitivity of the ICS to be 97.1% and 86.3%, respectively. There was excellent agreement between the results obtained by ELISA and the ICS (kappa = 0.843). Additionally, there was strong agreement between the results of bacterial isolation from pig tonsils and ICS test (kappa = 0.658). Because it is rapid and easy to use, the test is suitable for the serological surveillance of SS2 at farms.

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Validation of a Novel Commercial ELISA Test for the Detection of Antibodies against Coxiella burnetii

Pathogens ◽

10.3390/pathogens9121075 ◽

2020 ◽

Vol 9 (12) ◽

pp. 1075

Author(s):

Salvatore Ledda ◽

Cinzia Santucciu ◽

Valentina Chisu ◽

Giovanna Masala

Keyword(s):

Diagnostic Accuracy ◽

Phase Ii ◽

Coxiella Burnetii ◽

Q Fever ◽

Animal Health ◽

Elisa Test ◽

Clinical Diagnostics ◽

Complex Life Cycle ◽

Enzyme Linked Immunosorbent Assay ◽

Specificity And Sensitivity

Q fever is a zoonosis caused by Coxiella burnetii, a Gram-negative pathogen with a complex life cycle and a high impact on public and animal health all over the world. The symptoms are indistinguishable from those belonging to other diseases, and the disease could be symptomless. For these reasons, reliable laboratory tests are essential for an accurate diagnosis. The aim of this study was to validate a novel enzyme-linked immunosorbent assay (ELISA) test, named the Chorus Q Fever Phase II IgG and IgM Kit (DIESSE, Diagnostica Senese S.p.A), which is performed by an instrument named Chorus, a new device in medical diagnostics. This diagnostic test is employed for the detection of antibodies against C. burnetii Phase II antigens in acute disease. Our validation protocol was performed according to the Italian Accreditation Body (ACCREDIA) (Regulation UNI CEI EN ISO/IEC 17025:2018 and 17043:2010), OIE (World Organization for Animal Health), and Statement for Reporting Studies of Diagnostic Accuracy (STARD). Operator performance was evaluated along with the analytical specificity and sensitivity (ASp and ASe) and diagnostic accuracy of the kit, with parameters such as diagnostic specificity and sensitivity (DSp and DSe) and positive and negative predictive values (PPV and NPV), in addition to the repeatability. According to the evaluated parameters, the diagnostic ELISA test was shown to be suitable for validation and commercialization as a screening method in human sera and a valid support for clinical diagnostics.

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Effect of Antigen Coating Conditions on Enzyme-Linked Immunosorbent Assay for Detection of Immunoglobulin G Antibody to Neisseria meningitidis Serogroup Y and W135 Capsular Polysaccharide Antigens in Serum

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.10.6.1136-1140.2003 ◽

2003 ◽

Vol 10 (6) ◽

pp. 1136-1140 ◽

Cited By ~ 4

Author(s):

Peter C. Giardina ◽

Renee E. Evans ◽

Daniel J. Sikkema ◽

Dace Madore ◽

Stephen W. Hildreth

Keyword(s):

Immunoglobulin G ◽

Capsular Polysaccharide ◽

Enzyme Linked Immunosorbent Assay ◽

Meningococcal Vaccine ◽

Igg Antibody ◽

Human Sera ◽

Polysaccharide Antigens ◽

Reference Serum ◽

Adult Volunteers ◽

Assay Conditions

ABSTRACT Human sera collected from 28 consenting adult volunteers were used to define assay conditions for meningococcal vaccine clinical trial serology. Immunoassay parameters were optimized with these test sera and the standard reference serum, CDC1992. Coating conditions for serogroup Y and W135 polysaccharide antigens were found to influence the predicted serum immunoglobulin G (IgG) antibody concentrations. Sera that displayed IgG antibody binding profiles most unlike that of CDC1992 were influenced the most by coating conditions. Our results suggest that presentation of specific epitopes is influenced by antigen-coating concentrations for serogroup Y and W135 polysaccharides.

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Use of a Standardized Bovine Serum Panel To Evaluate a Multiplexed Nonstructural Protein Antibody Assay for Serological Surveillance of Foot-and-Mouth Disease

Clinical and Vaccine Immunology ◽

10.1128/cvi.00227-07 ◽

2007 ◽

Vol 14 (11) ◽

pp. 1472-1482 ◽

Cited By ~ 11

Author(s):

Julie Perkins ◽

Satya Parida ◽

Alfonso Clavijo

Keyword(s):

Nonstructural Protein ◽

Foot And Mouth Disease ◽

Mouth Disease ◽

Enzyme Linked Immunosorbent Assay ◽

Reference Laboratory ◽

Additional Information ◽

Mouth Disease Virus ◽

Foot And Mouth ◽

Antibody Assays ◽

Serological Surveillance

ABSTRACT Liquid array technology has previously been used to show proof of principle of a multiplexed nonstructural protein serological assay to differentiate foot-and-mouth disease virus-infected and vaccinated animals. The current multiplexed assay consists of synthetically produced peptide signatures 3A, 3B, and 3D and the recombinant protein signature 3ABC in combination with four controls. To determine the diagnostic specificity of each signature in the multiplex, the assay was evaluated against a naive population (n = 104) and a vaccinated population (n = 94). Subsequently, the multiplexed assay was assessed by using a panel of bovine sera generated by the World Reference Laboratory for foot-and-mouth disease in Pirbright, United Kingdom. This serum panel has been used to assess the performance of other singleplex enzyme-linked immunosorbent assay (ELISA)-based nonstructural protein antibody assays. The 3ABC signature in the multiplexed assay showed performance comparable to that of a commercially available nonstructural protein 3ABC ELISA (Cedi test), and additional information pertaining to the relative diagnostic sensitivity of each signature in the multiplex was acquired in one experiment. The encouraging results of the evaluation of the multiplexed assay against a panel of diagnostically relevant samples promote further assay development and optimization to generate an assay for routine use in foot-and-mouth disease serological surveillance.

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Multicolor enzyme-linked immunoassay method for visual readout of carbendazim

Analytical Methods ◽

10.1039/d1ay01028j ◽

2021 ◽

Author(s):

Haoran Liu ◽

Yiwen Wang ◽

Ruijie Fu ◽

Jing Zhou ◽

Yanlin Liu ◽

...

Keyword(s):

Immunosorbent Assay ◽

High Specificity ◽

Enzyme Linked Immunosorbent Assay ◽

Specificity And Sensitivity

Enzyme-linked immunosorbent assay (ELISA) with high specificity and sensitivity is one of the most popular techniques for detecting carbendazim (CBD), a commonly used benzimidazole fungicide in agriculture. However, the traditional...

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Study of Antibody Levels in Children with Purulent Otitis Media

Annals of Otology Rhinology & Laryngology ◽

10.1177/00034894800890s331 ◽

1980 ◽

Vol 89 (3_suppl) ◽

pp. 117-120 ◽

Cited By ~ 2

Author(s):

P. Branefors ◽

T. Dahlberg ◽

O. Nylén

Keyword(s):

Otitis Media ◽

Serum Antibody ◽

Acute Otitis Media ◽

High Serum ◽

Haemophilus Influenzae Type ◽

Enzyme Linked Immunosorbent Assay ◽

Polysaccharide Antigens ◽

Antibody Titers ◽

Iga Antibody ◽

Antibody Levels

A series of episodes of acute otitis media were studied with reference to the bacterial findings in the nasopharynx and the specific antibody response in a group of children nine months to ten years of age, with previous frequent episodes of acute otitis media, Serum IgG, IgM and IgA antibody levels against five polysaccharide antigens, namely Haemophilus influenzae type b and Streptococcus pneumoniae types 3, 6, 19 and 23, were studied by means of an enzyme-linked immunosorbent assay. The selection of polysaccharide antigens was based on isolation frequency. The sera to be tested were tenfold serially diluted. An extinction of 0.2 over the base was taken as the end-point titer and expressed as in-log10. The results showed that most children including those under three years of age showed increasing homologous antibody titers at an infection, or had already initially very high antibody titers, especially of the IgG class. The titers reached levels of 104 to 105. In some cases, however, it could be shown that high serum antibody titers did not give protection against a new infection with the same serological type of bacteria. It was also demonstrated that most children, regardless of age, had IgG and IgM titers against the heterologous antigens. In some cases the levels were quite high (103 to 104). However, the IgA antibody levels were lower and in a considerable number of samples antibodies were not even detectable.

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LOCATE Enzyme-Linked Immunosorbent Assay for Detection of Salmonella in Food: Collaborative Study

Journal of AOAC International ◽

10.1093/jaoac/81.2.419 ◽

1998 ◽

Vol 81 (2) ◽

pp. 419-437 ◽

Cited By ~ 4

Author(s):

Vidhya Gangar ◽

Michael S Curiale ◽

Armando D'onorio ◽

Carol Donnelly ◽

Paul Dunnigan ◽

...

Keyword(s):

Collaborative Study ◽

Screening Method ◽

Culture Method ◽

Microtiter Plate ◽

Black Pepper ◽

Soy Flour ◽

Food Type ◽

Enzyme Linked Immunosorbent Assay ◽

Data Sets ◽

Whole Egg

abstract A collaborative study was performed in 27 laboratories to validate the enzyme-linked immunosorbent procedure LOCATE for rapid detection of Salmonella in foods. Results were read visually and with a microtiter plate reader. The LOCATE method was compared with the Bacteriological Analytical Manual (BAM)/AOAC INTERNATIONAL culture method for detecting Salmonella in 6 foods: milk chocolate, nonfat dry milk, dried whole egg, soy flour, ground black pepper, and ground raw turkey. Two foods—dried whole egg and black pepper—required repeat rounds because insufficient data sets were produced initially (AOAC INTERNATIONAL stipulates a minimum of 15 sets per food type). Each laboratory tested one or more of the 6 foods. A total of 1 439 samples were analyzed, and no significant differences (P <0.05) were observed between LOCATE with either visual or reader detection and BAM/AOAC INTERNATIONAL results. The LOCATE screening method with visual or reader detection is recommended for Official First Action Approval

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Competitive Direct Enzyme-Linked Immunosorbent Assay for Detection of Sulfamethazine Residues in Swine Urine and Muscle Tissue

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/71.6.1137 ◽

1988 ◽

Vol 71 (6) ◽

pp. 1137-1140 ◽

Cited By ~ 2

Author(s):

Deborah E Dixon-Holland ◽

Stanley E Katz

Keyword(s):

Horseradish Peroxidase ◽

Muscle Tissue ◽

Immunosorbent Assay ◽

Microtiter Plate ◽

Test System ◽

Enzyme Linked Immunosorbent Assay ◽

Sensitive Assay ◽

Saline Extract ◽

Swine Urine ◽

Phosphate Buffered Saline

Abstract A sensitive assay for the detection of sulfamethazine in swine urine and muscle tissue using a direct competitive enzyme-linked immunosorbent assay (ELISA) has been developed. Undiluted urine or a phosphate-buffered saline extract of pork muscle tissue is mixed with an enzyme-labeled conjugate of sulfamethazine and horseradish peroxidase. The mixture is added to wells of a microtiter plate coated with antibody to sulfamethazine. After the test system is incubated, washed, and re-incubated with substrate and the reaction is stopped, the absorbance is measured at 405 nm. Levels of sulfamethazine as low as 20 ng sulfamethazine/g muscle tissue and 10 ng sulfamethazine/ mL swine urine were detected and estimated

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The development and application of an indirect Elisa test for the detection of antibodies to bovine respiratory syncytial virus in blood serum

Veterinární Medicína ◽

10.17221/7848-vetmed ◽

2001 ◽

Vol 46 (No. 2) ◽

pp. 29-34 ◽

Cited By ~ 5

Author(s):

K. Kovařčík

Keyword(s):

Respiratory Syncytial Virus ◽

Neutralization Test ◽

Elisa Test ◽

Bovine Respiratory Syncytial Virus ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Neutralization ◽

Specificity And Sensitivity ◽

Reference Serum ◽

Serum Neutralization Test ◽

Syncytial Virus

We developed an indirect enzyme-linked immunosorbent assay (ELISA) for detection of serum antibodies to bovine respiratory syncytial virus. For evaluation of the newly developed ELISA, field sera collected from 549 head of cattle in the Czech Republic were tested in parallel by a serum neutralization test. The tests showed 98.36% agreement. The specificity and sensitivity of the ELISA relative to serum neutralization test was 97.00% (226/233) and 99.37% (314/316), respectively. Tissue culture-grown viral antigen was used in the tests. The corrected optical density (COD) of each sample tested at dilution 1/100 was expressed as a percentage of the COD of a positive reference serum included on each plate, this value was the sample/positive (S/P) ratio. We determined the relationship between the S/P ratio (%) obtained at a dilution 1/100 and the end point titer calculated by serum neutralization test (r = 0.9743). The ELISA test was evaluated by testing acute and convalescent (3 wk later) serum pairs from 9 head of cattle with confirmed BRSV infection for demonstration of seroconversion. The ELISA test demonstrated a clear increase of the S/P ratio (%) between acute and convalescent serum pairs (on average 42.2 ± 13.1).

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