Establishment of an Indirect ELISA Method for the Detection of the Bovine Rotavirus VP6 Protein
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Cell and Serum Samples
2.2. Construction and Characterization of the VP6 Recombinant Plasmid
2.3. Transfection, Purification, and Western Blot Analysis of Recombinant Proteins
2.4. Preparation of Mouse Polyclonal Antibodies
2.5. Establishment of an Indirect ELISA Based on the VP6 Protein
2.6. Determination of Critical Values
2.7. Sensitivity and Specificity Tests
2.8. Repeat Experiments within and between Batches
2.9. Preliminary Application of ELISA in Clinical Samples
3. Results
3.1. Enzymatic Digestion of Recombinant Plasmids and Protein Expression
3.2. Establishment of an Indirect ELISA Method Based on a Polyclonal Antibody against the VP6 Protein
3.3. Determination of Critical Values
3.4. Validation of the Sensitivity and Specificity of an Indirect ELISA Based on a Murine Polyclonal Antibody against the VP6 Protein
3.5. Stability Verification
3.6. Preliminary Application of ELISA in Clinical Samples
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Batch | VP6 Mouse Polyclonal Antibody | Average OD450 nm | Standard Deviation SD | Coefficient of Variation/% CV |
---|---|---|---|---|
Within batch | Immune VP6 mouse polyclonal antibody | 0.600 | 0.033 | 5.5% |
Negative mouse antibody | 0.233 | 0.006 | 2.6% | |
Between batches | Immune VP6 mouse polyclonal antibody | 0.525 | 0.045 | 8.6% |
Negative mouse antibody | 0.219 | 0.012 | 5.5% |
BRV Positive Samples | BRV Negative Sample | ||
---|---|---|---|
iELISA Positive | iELISA Negative | iELISA Positive | iELISA Negative |
21 | 3 | 0 | 40 |
Sensitivity = [21/21 + 3] = 87.5% | |||
Specificity = [40/40] = 100% |
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Niu, X.; Liu, Q.; Wang, P.; Zhang, G.; Jiang, L.; Zhang, S.; Zeng, J.; Yu, Y.; Wang, Y.; Li, Y. Establishment of an Indirect ELISA Method for the Detection of the Bovine Rotavirus VP6 Protein. Animals 2024, 14, 271. https://doi.org/10.3390/ani14020271
Niu X, Liu Q, Wang P, Zhang G, Jiang L, Zhang S, Zeng J, Yu Y, Wang Y, Li Y. Establishment of an Indirect ELISA Method for the Detection of the Bovine Rotavirus VP6 Protein. Animals. 2024; 14(2):271. https://doi.org/10.3390/ani14020271
Chicago/Turabian StyleNiu, Xiaoxia, Qiang Liu, Pu Wang, Gang Zhang, Lingling Jiang, Sinong Zhang, Jin Zeng, Yongtao Yu, Yujiong Wang, and Yong Li. 2024. "Establishment of an Indirect ELISA Method for the Detection of the Bovine Rotavirus VP6 Protein" Animals 14, no. 2: 271. https://doi.org/10.3390/ani14020271
APA StyleNiu, X., Liu, Q., Wang, P., Zhang, G., Jiang, L., Zhang, S., Zeng, J., Yu, Y., Wang, Y., & Li, Y. (2024). Establishment of an Indirect ELISA Method for the Detection of the Bovine Rotavirus VP6 Protein. Animals, 14(2), 271. https://doi.org/10.3390/ani14020271