Evaluation of Antibodies Produced against Lipopolysaccharides of Vibrio cholerae for Rapid Detection of Cholera

Authors
M. R. Akbari 1
A. Ahmadi 2
J. Salimian 3
1 Department of Biology, Faculty of Basic Science, IHU University, Tehran, Iran
2 Molecular Biology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
3 Chemical Injuries Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
Abstract
Objective: Cholera is an endemic disease in Iran. Early detection, especially in times of disease outbreaks, is of vital importance. The antibody against the lipopolysaccharide (LPS) is an important method for bacterial detection. This study intends to extract and purify the LPS of Vibrio cholerae and evaluate the cholera antibody for detection purposes.
Methods: Vibrio cholerae was cultured in tryptone extract medium. LPS was extracted by the hot phenol water method, purified, and dialyzed. We measured the LPS protein and sugar content, purity, and biological activity. Antibodies were produced by injection of the killed bacteria with Freund's complete adjuvant into rabbits and then the LPS was injected three times with Freund's incomplete adjuvant. After the last booster, blood samples were taken. We used ELISA to determine the antibody titers against the Inaba and Ogawa serotypes, the LPS of these serotypes, and several other similar bacteria.
Results: The amount of protein in the purified LPS was approximately zero and sugar was 0.5 mg/ml. The LPS had a titer activity of 1024, and consisted of three bands (5.2, 4, and 5.14 KD). Antibodies produced by the rabbits identified the bacterial Inaba and Ogawa serotypes, and the purified LPS. Ogawa and Inaba serotypes cross-react with each other but not with other species of Vibrio and other bacteria. The LPS antibody titer against the Ogawa serotype was 1:32000, whereas for Inaba it was 1:16000.
Conclusion: Due to the low cost of production, high sensitivity, and importance of cholera diagnosis in Iran, the antiserum produced in this study can be used as a tool for early screening of cholera and discrimination of O1 strains from non-O1 strains in immunologic tests.

Keywords

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Pathobiology Reserach
Volume 18, Issue 4
January 2016
Pages 23-31