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Journal of Applied Sciences and Environmental Management
World Bank assisted National Agricultural Research Project (NARP) - University of Port Harcourt
ISSN: 1119-8362
Vol. 9, No. 1, 2005, pp. 5-10
Bioline Code: ja05001
Full paper language: English
Document type: Research Article
Document available free of charge

Journal of Applied Sciences and Environmental Management, Vol. 9, No. 1, 2005, pp. 5-10

 en Molecular analysis of cross-bacterial contamination detected in biotin-free buffers during diagnosis of HCVinfections
EL-RASHDY M. REDWAN; DESOUKY ABD-EL-HALEEM

Abstract

In a routine work to determine Hepatitis C Virus (HCV) molecules in human infected serum with biotin/streptavidine enzyme linked immunosorbant assay (ELISA) technique, unexpected false positive was observed. No positive signals were noticed after changing all ELISA buffers. Subsequently, contaminated buffers were screened and analyzed for microbial contamination. Out of fifty-five, five randomly selected bacterial colonies were examined for biosynthesis of biotin using ELISA and/or Western blot binding biotin techniques in both supernatants and cell pellets. In compare to the E.coli reference strain a strong biotin signals in all examined isolates were recorded. All isolates were then examined for their genetic heterogeneity by PCR-RFLP technique of 23S rDNA genes. While, isolate BP(R2) which gave the highest biotin signal, was subjected for molecular identification. Comparative sequence analysis of the 16S rDNA gene (~1440 bp) revealed that this isolate is a member of the bacterial genus Delftia exhibiting a similarity value of 99.3% with Delftia acidovorans check for this species in other resources . In conclusion, a soluble biotin is secreted by the isolate Delftia acidovorans BP(R2) and it is also coupled to protein with molecular weight 25-26 KDa.As well as, this bacterial contaminationwas the reason for the false positive results observed during the detection of HCV infections. @JASEM

 
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