title: Hybridoma as a specific, sensitive, and ready to use sensing element: a rapid fluorescence assay for detection of Vibrio cholerae O1.
abstract: Over the last decade, isolation and purification of monoclonal antibodies, for diagnostic analysis, have been carried out using the hybridoma expression system. The present study describes a novel example of a detection system using hybridoma cells containing antibody against O1 antigen directly for V. cholerae diagnosis, which is a major health problem in many parts of the world, especially in developing countries. This method has advantages such as simplicity, ease of process, and it does not require manipulation of hybridoma cell. For this approach, an efficient amount of fluorescence calcium indicator, fura 2-AM, was utilized, which emitted light when the intracellular calcium concentration increased as result of antigen binding to specific antibody. More reliable results are obtained via this method and it is considerably faster than other methods, which has the response time of less than 45 s for detection of V. Cholerae O1. Also, the limit of detection was computed to be 50 CFU/mL (<13 CFU per assay). In addition, no significant responses were observed in the presence of other bacteria with specific hybridoma or other cell lines exposed to V. cholerae O1. Furthermore, this method was successfully applied to V. cholerae O1 detection in spiked environmental samples, including water and stool samples without any pretreatment. All results reveal that hybridoma cells can provide a valuable, simple, and ready to use tool for rapid detection of other pathogenic bacteria, toxins, and analytes.
authors.affiliation: Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: Parichehr
authors.initials: P
authors.lastname: Zamani
authors.affiliation: Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran. sajedi_r@modares.ac.ir.
authors.forename: Reza H
authors.initials: RH
authors.lastname: Sajedi
authors.affiliation: Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: Saman
authors.initials: S
authors.lastname: Hosseinkhani
authors.affiliation: Department of Bioscience and Biotechnology, Mallek Ashtar University of Technology, Tehran, Iran.
authors.forename: Mehdi
authors.initials: M
authors.lastname: Zeinoddini
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002118:Calcium / Q000032:analysis; D065096:Calcium Chelating Agents / Q000737:chemistry; D002771:Cholera / Q000382:microbiology; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry*; D016257:Fura-2 / Q000737:chemistry*; D006825:Hybridomas; D057230:Limit of Detection; D051379:Mice; D013050:Spectrometry, Fluorescence / Q000379:methods*; D041241:Vibrio cholerae O1 / Q000302:isolation & purification*; D014871:Water Microbiology
chemical_list: D000911:Antibodies, Monoclonal; D065096:Calcium Chelating Agents; D005456:Fluorescent Dyes; D002118:Calcium; D016257:Fura-2
keywords: Calcium increasing; Fluorescence emission; Fura 2-AM; Hybridoma; Monoclonal antibody; Vibrio cholerae O1