Development of Highly Specific Monoclonal Antibodies for the Diagnosis ofVibrio cholerae01

Abstract

We report here the development of two monoclonal antibodies, termed 5G8 and 5C12, belonging to the IgM and IgG1 class, respectively, suitable for the identification of Vibrio cholerae 01 in clinical and environmental samples. The specificities of the monoclonals were evaluated by ELISA and indirect immunofluorescent microscopy of microorganisms normally present in stool samples and with two bacterial panels. One panel included 72 potentially antigenically related bacterial strains and the second panel included 20 pathogenic bacterial strains involved in diarrhea cases. The results of these extensive analyses indicate that monoclonal antibodies 5G8 and 5C12 are highly specific and suitable for the clinical diagnosis of Vibrio cholerae 01 in human stool samples by indirect immunofluorescent microscopy. Although the antigenic sites recognized by these antibodies were not identified in this study, the observation of Western blot patterns suggested that 5G8 and 5C12 monoclonal antibodies bind to LPS epitopes, a good structural marker for the detection of V. cholerae 01 because it is present in all bacterial cell walls.

Characteristics of monoclonal antibodies against Piscirickettsia salmonis

“…2b). The control blot carried out with the monoclonal antibody 5C12 against V. cholerae (Castillo et al. 1995) did not cross‐react with any antigen of P. salmonis (Fig.…”

“…Two‐month‐old female BALB/c mice were injected intraperitoneally at 1 week intervals four times with P. salmonis LF‐89 or the Bios Chile or ISP isolates (10 7 cfu per mouse) resuspended in PBS (Castillo et al. 1995).…”

“…1979). With each blot analysis a strip was incubated with the irrelevant monoclonal antibody 5C12 against V. cholerae (Castillo et al. 1995).…”

Vibrio cholerae O1 Strain TSI-4 Produces the Exopolysaccharide Materials That Determine Colony Morphology, Stress Resistance, and Biofilm Formation

“…For immunoelectron microscopy, a colloidal gold probe (Wako Pure Chemical Industries Ltd., Osaka, Japan) was used to label the specific reaction sites of anti-EPS serum and an anti-O1 monoclonal antibody (mouse immunoglobulin G anti-V. cholerae monoclonal antibody reacting with lipopolysaccharide [LPS] epitopes present in the bacterial cell wall). The anti-V. cholerae monoclonal antibody was purchased from Cosmo Bio Co., Ltd. (6). To label the specimens, 1 ml of a bacterial suspension of about 10 8 CFU/ml was treated with antiserum appropriately diluted with phosphate-buffered saline (PBS) for 30 min at 37°C.…”

Proteolytic activity of rabbit perivitelline spermatozoa

“…Group c: Spermatozoa from 10 rabbit zygotes were used to inseminate 17 freshly ovulated rabbit eggs in the presence of 600 µg/ml of the monoclonal antibody 5C12 specific to Vibrio cholerae 01 to assess the specificity of the inhibition. This is a negative control in the in vitro fertilisation assay, because the anti-V. cholerae 01 antibody belongs to the same IgG subclass as the antiacrosin antibody and is directed against LPS (Castillo et al, 1995). Thus it is irrelevant for the spermatozoa…”