Highly sensitive Escherichia coli O157:H7 detection in a large volume sample using a conical polymer tube chamber consisting of micro-glass beads
| Autor: | Ji Yeong Won, Junhong Min |
|---|---|
| Rok vydání: | 2010 |
| Předmět: |
Cell Culture Techniques
Biomedical Engineering Biophysics Biosensing Techniques Escherichia coli O157 medicine.disease_cause Buffer (optical fiber) Specimen Handling Adsorption Molecular beacon Electrochemistry medicine Water Pollutants Escherichia coli chemistry.chemical_classification Chromatography Chemistry Equipment Design General Medicine Polymer NASBA Bacterial Load Microspheres Equipment Failure Analysis Biochemistry Sample Size Nucleic acid Glass Water Microbiology Molecular probe Biotechnology |
| Zdroj: | Biosensors and Bioelectronics. 26:112-117 |
| ISSN: | 0956-5663 |
| DOI: | 10.1016/j.bios.2010.05.025 |
| Popis: | In this study, we developed a method for the highly sensitive detection of viable pathogenic bacteria in a large volume sample by performing RNA concentration, amplification, and using fluorescently tagged capture probes in a single polymer chamber without transferring RNA samples from one chamber to another. Nucleic acids were extracted from Escherichia coli O157:H7 and loaded into glass micro-beads embedded in a conical polymer tube chamber. Nucleic acids were concentrated in the micro-tube in a low pH buffer (pH 5). The mRNA, which was adsorbed to the glass micro-beads, was amplified by Nucleic Acid Sequence Based Amplification in the same chamber at a relatively high pH (pH 8.9). The products amplified were measured using the hair-loop type detection probe with FAM and DABCYL, which was pre-mixed in the NASBA mater mixture. As a result, high sensitivity (100% for rain water and 60% for river water) with less than 10 viable E. coli O157:H7 in 100 ml could be achieved within 4 h using the simple method proposed in this study. |
| Databáze: | OpenAIRE |
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