Enzyme-linked immunosorbent assay for detecting anti-pertussis toxin antibody in mouse.
- Author:
Gi Sub CHOI
1
;
Dong Ho HUH
;
Seung Beom HAN
;
Dong Ho AHN
;
Kyu Ri KANG
;
Ji Ahn KIM
;
Bo Mi CHOI
;
Hea Ryun KIM
;
Jin Han KANG
Author Information
- Publication Type:Original Article
- Keywords: Bordetella pertussis; Enzyme-linked immunosorbent assay; Murine; Whooping cough
- MeSH: Animals; Antibodies; Bordetella pertussis; Chromatography; Diagnosis; Enzyme-Linked Immunosorbent Assay*; Horseradish Peroxidase; Methods; Mice*; Models, Animal; Pertussis Toxin; Streptavidin; Vaccines; Whooping Cough
- From:Clinical and Experimental Vaccine Research 2019;8(1):64-69
- CountryRepublic of Korea
- Language:English
- Abstract: PURPOSE: Although the DTaP and Tdap vaccines used to prevent pertussis have been used for a long time, there is no standard method for measuring pertussis antigens. Therefore, this preliminary study was conducted to develop an enzyme-linked immunosorbent assay method using an animal model for measuring antibodies against pertussis toxin, the most important pertussis pathogenic antigen, in the sera of vaccinated mice. MATERIALS AND METHODS: Bordetella pertussis Tohama phase I was cultured for 24–30 hours, and then pertussis toxin was purified from the culture medium by chromatography. Purified pertussis toxin was diluted in phosphate-buffered saline-coating buffer, and 100 µL of diluted pertussis toxin was added to each well and reacted at room temperature for 4 hours. Positive serum was diluted to 1/1,250–1/80,000 and negative serum was diluted to 1/50 to determine the coating concentration with the optimal signal/noise ratio. Optimal test conditions were confirmed from the dilution factors of the secondary antibody and streptavidin horseradish peroxidase (SA-HRP). RESULTS: Optimal conditions were as follows: 4 µg/mL for coating antigen; 1/40,000 for secondary antibody; and 1/1,000 for the SA-HRP dilution factor. Comparison of the sera obtained from mice treated with a developing vaccine and commercial vaccine with National Institute for Biological Standard and Control standard serum under the established conditions showed the following results: 1,300.62, 534.94, and 34.85, respectively. CONCLUSION: The method developed in this study is suitable for measuring anti-pertussis toxin antibodies and may be applicable for clinical sample analysis or indirect diagnosis of pertussis.
