Abstract
An antigen capture enzyme-linked immunosorbent assay (AC-ELISA) was established based on two monoclonal antibodies (mAbs) for the quantification of equine infectious anemia virus (EIAV). Two p26-specific monoclonal antibodies were developed in mice. The mAb 9H8 was coated in microtiter plates as the capture antibody; the other mAb, 1G11, was coupled to horseradish peroxidase (HRP) and used as the detection antibody. The limit of detection for the EIAV p26 protein was 0.98 ng/ml, and the linearity range was 3.9–62.5 ng/ml. The sensitivity of p26 AC-ELISA for the detection of the virus (EIAV infectious clone, FDDVcmv3–8) was the same as that for the purified p26 protein. No cross-reaction with other equine viruses was observed by this method. The intra- and inter-assay coefficients of variation were below 8.3 and 10.3 % for testing p26 and FDDVcmv3–8, respectively. The AC-ELISA was also compared to Western blotting (WB) and reverse transcriptase (RT) assays, validating the sensitivity, accuracy, and reliability of this method. Both the AC-ELISA and RT assay showed good agreement, with a correlation coefficient of R 2 = 0.9946. Sample analysis showed that this AC-ELISA is a useful tool for quantifying EIAV p26 in cell lysates and culture medium.
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Acknowledgments
This study was supported by grants from the National Natural Science Foundation of China (31222054 to **aojun Wang and 31300713 to Zhe Hu) and grants from the State Key Laboratory of Veterinary Biotechnology (SKLVBP201426 to Zhe Hu).
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Hu, Z., Chang, H., Ge, M. et al. Development of antigen capture ELISA for the quantification of EIAV p26 protein. Appl Microbiol Biotechnol 98, 9073–9081 (2014). https://doi.org/10.1007/s00253-014-6078-8
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DOI: https://doi.org/10.1007/s00253-014-6078-8