Development of competitive ELISA for neosporosis by employing immunoproteomics

Authors: Shin, Yong-seung¹; Lee, Eung-goo²; Shin, Gee-wook¹; Kim, Young-rim¹; Lee, Eun-young¹; Kim, Jae-hoon³; Jang, Hwan⁴; Kim, Dae-yong⁵; Kim, Yong-hwan¹; Kim, Gon-sup¹; Suh, Myung-deuk¹; Jung, Tae-sung⁶

Source: Clinical Proteomics, Volume 1, Numbers 3-4, September 2004 , pp. 235-248(14)

Publisher: Humana Press

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Abstract:

In this study, proteomics was used to explore the antigenic proteins that are involved in cross-reactivity during serodiagnosis between Neospora caninum (N. caninum) and Toxoplasma gondii (T. gondii). Competitive enzyme-linked immunosorbent assay (C-ELISA) developed by proteomics shed a new light on the infection of N. caninum. Cross-reactivity of antigenic proteins between N. caninum and T. gondii tachyzoites was explored by using the conventional sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) (1-DE) and two-dimensional gel electrophoresis (2-DE) immunoblot. The proteins were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. The protein expression patterns in the immunoblot profiles of N. caninum were similar to bovine, chicken, and rabbit anti-N. caninum serum, but they were not similar to rabbit anti-T. gondii serum. Band at 79 kDa, HSP70, and actin on immunoblot profiles reacted, in general, with bovine, chicken, and rabbit anti-N. caninum serum and also with rabbit anti-T. gondii serum, respectively. Whereas the band at 144 kDa, and NCDG-1 were detected on bovine, chicken, and rabbit anti-N. caninum immunoblot profiles, they were not observed on rabbit anti-T. gondii immunoblot profile. These specific antigenic proteins were recorded as species-specific proteins of N. caninum against T. gondii. Based on the proteome analysis, C-ELISA was developed to screen the cattle infected with N. caninum by using N. caninum tachyzoite lysate as a coating antigen and chicken anti-N. caninum immunoglobulin (Ig)Y as a competitor. C-ELISA was able to detect the antibody of N. caninum without cross-reactivity with T. gondii. Furthermore, it achieved a fine diagnostic performance in the cases of 162 bovine sera.

Keywords: Neospora caninum; Toxoplasma gondii; cross-reactivity; competitive ELISA; immunoproteomics

Document Type: Research article

DOI: 10.1385/CP:1:3-4:235

Affiliations: 1: Institute of Animal Medicine, College of Veterinary Medicine, Gyeongsang National University, 900 Gazwa-Dong, 660-701, Jinju, Gyeong-nam, Republic of Korea, 2: Department of Molecular Parasitology, School of Medicine, Sungkyunkwan University Suwon, Republic of Korea, 3: Department of Veterinary Medicine, Cheju National University, Jeju, Republic of Korea, 4: National Veterinary Research and Quarantine Service, Anyang, Republic of Korea, 5: Department of Veterinary Pathology, College of Veterinary Medicine, Seoul National University, Seoul, Republic of Korea, 6: Institute of Animal Medicine, College of Veterinary Medicine, Gyeongsang National University, 900 Gazwa-Dong, 660-701, Jinju, Gyeong-nam, Republic of Korea, Email: jungts@nongae.gsnu.ac.kr

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