Comparative evaluation of indirect-ELISA and DOT blot assay for serodetection of Mycoplasma gallisepticum and Mycoplasma synoviae antibodies in poultry

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.

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Development of an Enzyme-Linked Immunosorbent Assay Using a Recombinant LigA Fragment Comprising Repeat Domains 4 to 7.5 as an Antigen for Diagnosis of Equine Leptospirosis

Clinical and Vaccine Immunology ◽

10.1128/cvi.00245-13 ◽

2013 ◽

Vol 20 (8) ◽

pp. 1143-1149 ◽

Cited By ~ 10

Author(s):

Weiwei Yan ◽

Muhammad Hassan Saleem ◽

Patrick McDonough ◽

Sean P. McDonough ◽

Thomas J. Divers ◽

...

Keyword(s):

Immunosorbent Assay ◽

Indirect Elisa ◽

Enzyme Linked Immunosorbent Assay ◽

Dot Blot ◽

Serum Dilution ◽

Content Type ◽

Diagnostic Antigen ◽

Repeat Domain ◽

Associated Proteins ◽

Dot Blot Assay

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.

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Development of an Indirect ELISA and Dot-Blot Assay for Serological Detection of Rice Tungro Disease

BioMed Research International ◽

10.1155/2017/3608042 ◽

2017 ◽

Vol 2017 ◽

pp. 1-7 ◽

Cited By ~ 1

Author(s):

Magdline Sia Henry Sum ◽

Siew Fung Yee ◽

Lily Eng ◽

Evenni Poili ◽

Julia Lamdin

Keyword(s):

Sensitivity And Specificity ◽

Rural Areas ◽

Indirect Elisa ◽

High Sensitivity ◽

Visual Observation ◽

Alternative Methods ◽

Dot Blot ◽

Specificity And Sensitivity ◽

Rice Tungro Disease ◽

Dot Blot Assay

Rice tungro disease (RTD) is one of the most destructive diseases of rice in South and Southeast Asia. RTD is routinely detected based on visual observation of the plant. However, it is not always easy to identify the disease in the field as it is often confused with other diseases or physiological disorders. Here we report the development of two serological based assays for ease of detection of RTD. In this study we had developed and optimized an indirect ELISA and dot-blot assay for detection of RTD. The efficiency of both assays was evaluated by comparing the specificity and sensitivity of the assays to PCR assay using established primer sets. The indirect ELISA showed 97.5% and 96.6%, while the dot-blot assay showed 97.5% and 86.4% sensitivity and specificity, respectively, when compared to established PCR method. The high sensitivity and specificity of the two assays merit the use of both assays as alternative methods to diagnose RTD. Furthermore, the dot-blot assay is a simple, robust, and rapid diagnostic assay that is suitable for field test for it does not require any specialized equipment. This is a great advantage for diagnosing RTD in paddy fields, especially in the rural areas.

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APPLICATION OF REAL-TIME PRC TECHNIQUE AND REVERSE DOT-BLOT FOR DETECTION THE HIGH RISK TYPES OF HUMAN PAPILLOMAVIRUS CAUSING CERVICAL CANCER

Journal of Medicine and Pharmacy ◽

10.34071/jmp.2017.3.15 ◽

2017 ◽

pp. 99-103

Author(s):

Van Bao Thang Phan ◽

Hoang Bach Nguyen ◽

Van Thanh Nguyen ◽

Thi Nhu Hoa Tran ◽

Viet Quynh Tram Ngo

Keyword(s):

Cervical Cancer ◽

High Risk ◽

Real Time ◽

Real Time Pcr ◽

Dot Blot ◽

The Real ◽

Dot Blot Assay ◽

Hpv Types ◽

High Risk Hpv ◽

Reverse Dot Blot

Introduction: Infection with HPV is the main cause of cervical cancer. Determining HPV infection and the types of HPV plays an important role in diagnosis, treatment and prognosis of cervicitis/cervical cancer. Aims: Determining proportion of high-risk HPV types and the occurrence of coinfection with multiple HPV types. Methods: 177 women with cervicitis or abnormal Pap smear result were enrolled in the study. Performing the real-time PCR for detecting HPV and the reverse DOT-BLOT assay for determining type of HPV in cases of positive PCR. Results: 7 types of high-risk HPV was dectected, the majority of these types were HPV type 18 (74.6%) and HPV type 16 (37.6%); the proportion of infection with only one type of HPV was 30.4% and coinfection with multiple HPV types was higher (69.6%), the coinfected cases with 2 and 3 types were dominated (32.2% and 20.3%, respectively) and the coinfected cases with 4 and 5 types were rare. Conclusion: Use of the real-time PCR and reverse DOT-BLOT assay can determine the high-risk HPV types and the occurrence of coinfection with multiple HPV types. Key words: HPV type, Reverse DOT-BLOT, real-time PCR,PCR, cervical cancer

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Use of monoclonal and polyclonal antibodies to detect prostatic acid phosphatase by immunoblotting.

Clinical Chemistry ◽

10.1093/clinchem/32.10.1832 ◽

1986 ◽

Vol 32 (10) ◽

pp. 1832-1835 ◽

Cited By ~ 2

Author(s):

P C Patel ◽

L Aubin ◽

J Côte

Keyword(s):

Monoclonal Antibodies ◽

Acid Phosphatase ◽

Western Blot ◽

Polyclonal Antibodies ◽

Prostatic Acid Phosphatase ◽

The Other ◽

Dot Blot ◽

Western Blots ◽

Dot Blot Assay ◽

Polyclonal Antisera

Abstract We investigated two techniques of immunoblotting--the Western blot and the dot blot--for use in detecting prostatic acid phosphatase (PAP, EC 3.1.3.2). We used polyclonal antisera to human PAP, produced in rabbits by hyperimmunization with purified PAP, and PAP-specific monoclonal antibodies in the immunoenzymatic protocols. We conclude that PAP can be readily detected by Western blots with use of polyclonal antisera, but not with monoclonal antibodies. On the other hand, using a dot blot assay, we could easily detect PAP with both polyclonal and monoclonal antibodies.

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