Enrichment-ELISA for Detection of Salmonella typhi From Food and Water Samples

Key words: 
• Enrichment-ELISA /
• Food /
• Monoclonal antibody /
• Salmonella /
• Sandwich ELISA /
• Water

Abstract: Objective Development of monoclonal antibody based sandwich enzyme linked immunosorbant assay(sELISA)for rapid detection of Salmonella enterica serovar typhi(S.typhi) from food and water samples and optimization of enrichment procedures for use with the developed sELISA to increase the detection Sensitivity of the assay. Methods Spleen cells from BALB/c mice immunized with flagellin(H=d)antigen of S.typhi were fused with Sp2/0 myeloma cells.The hybridoma cell line specific to H=d antigen was established.characterized and ascites raised against one of these clones.The hypefimmune serum to flagellin antigen was raised in New Zealand White rabbits.An sELISA was developed using polyclonalantibody as capture and monoelonal antibody as detection antibody.To design the efficient culture sUrategies for use with the sELISA.different pre-enrichment and enrichment brothswere evaluated.The mediaincluded buffered peptonewater(BPW)and brain heart infusion broth for pre-enrichment and selenite F broth and Rappaport-Vassiliadis broth as enrichment broths.The developed sELISA with preceding enrichment step in BPW(Enrichment-ELISA)was evaluated in various food samples artificially inoculated with S. typhi bacteria.Various food(30)and water(35)samples collected from field were also tested by Enrichment-ELISA and culture method. Results Out of four specific clones to H=d antigen,one clone(#2/56.IgG2a isotype)was usedin sELISA.The sELISA had the detectionlimit of 104-105 cfu of S.typhi.Of the various broths used with sELISA,BPW was found to yield maximum ELISA values.Enrichment-ELISA,when tested in artificially inoculated food samples,generally,could detect 102S.typhi cfu/mL within 10 h from variousfood rinses(meat,vegetable)and milk samples.After overnight enrichment in BPW,as less as 2 bacteria per 10 mL of milk,meat rinse.and chicken rinse could be detected.Only one of the field samples(water)gave false positive result by Enrichment-ELISA.Conclusion In comparison to culture,the Enrchment-ELISA is a rapid.sensitive.and specific method for detection of S.typhi from food or water samples.This method may be used as rapid screening procedure for environmental monitoring during outbreak situation.