Development of Polyclonal Antibody against Clenbuterol for Immunoassay Application
Abstract
:1. Introduction
2. Results and Discussion
2.1. Polyclonal Antibody against CLB
2.2. Purification of Antibody
2.3. Characterization of Polyclonal Antibody
2.3.1. Antibody Titer Determination
2.3.2. Protein Gel Electrophoresis by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE)
2.3.3. Performance of the Polyclonal Anti-CLB Antibody Activities as Bio-Receptor
2.3.4. Cross-Reactivity
2.3.5. Recovery of CLB
3. Materials and Methods
3.1. Materials
3.2. Preparation of Immunization Solutions
3.3. Production of Polyclonal Anti-CLB Antibody
3.4. Purification of Antibody
3.5. Characterization of Polyclonal Antibody
3.5.1. Antibody Titer ELISA
3.5.2. Protein Gel Electrophoresis
3.5.3. Direct Competitive ELISA Analysis
3.5.4. Recovery Study
4. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
Abbreviations
CLB | clenbuterol |
BSA | bovine serum albumin |
ELISA | enzyme-linked immunosorbent assay |
FCA | Freund’s complete adjuvant |
FIA | Freund’s incomplete adjuvant |
HPLC | high-performance liquid chromatography |
HRP | Horseradish peroxidase |
Ig | Immunoglobulin |
LC-MS | liquid chromatography-mass spectrometry |
OVA | ovalbumin |
PBS | Phosphate buffer saline |
p-NPP | 4-nitrophenyl phosphate disodium salt hexahydrate |
SDS-PAGE | sulfate-polyacrylamide gel electrophoresis |
TEMED | 1,2-di-(dimethylamino)ethane |
TMB | 3,3′,5,5′–tetramethylbenzidine |
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Sample Availability: Samples of the compound polyclonal anti-CLB antibody is available from the authors. |
Family | Antibiotics | Structure |
---|---|---|
β-agonist | Clenbuterol | |
Salbutamol | ||
Mabuterol | ||
Ractopamine | ||
Terbutaline | ||
Other | Vancomycin | |
Tetracycline | ||
Chloramphenicol | ||
Streptomycin | ||
Nitrofuran |
Samples | Spiked Concentration (ng mL−1) | Average Recovery (ng mL−1) | Recovery (%) | Coefficient of Variation (%) |
---|---|---|---|---|
Milk A | 5 | 6.91 | 138 | 8.5 |
10 | 10.17 | 102 | 10.9 | |
Milk B | 5 | 5.76 | 115 | 3.9 |
10 | 9.24 | 92 | 6.9 |
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Talib, N.A.A.; Salam, F.; Sulaiman, Y. Development of Polyclonal Antibody against Clenbuterol for Immunoassay Application. Molecules 2018, 23, 789. https://doi.org/10.3390/molecules23040789
Talib NAA, Salam F, Sulaiman Y. Development of Polyclonal Antibody against Clenbuterol for Immunoassay Application. Molecules. 2018; 23(4):789. https://doi.org/10.3390/molecules23040789
Chicago/Turabian StyleTalib, Nurul Ain A., Faridah Salam, and Yusran Sulaiman. 2018. "Development of Polyclonal Antibody against Clenbuterol for Immunoassay Application" Molecules 23, no. 4: 789. https://doi.org/10.3390/molecules23040789