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Sun H.,Key Laboratory of Diarrhea Disease Detection | Mo Q.-H.,Key Laboratory of Diarrhea Disease Detection | Lin J.-C.,Key Laboratory of Diarrhea Disease Detection | Yang Z.,Key Laboratory of Diarrhea Disease Detection | And 4 more authors.
World Journal of Microbiology and Biotechnology | Year: 2011

In this study, we describe a DNA microarray assay by using bead-mediated visible light-assisted signal detection for simultaneous screening of seven clinically important enteric pathogens, including Escherichia coli O157:H7, Vibrio cholerae, Vibrio parahaemolyticus, Salmonella spp., Staphylococcus aureus, Rotavirus, and Norwalk virus (including genogroup I and II). Seven pairs of primers, in which the forward primers were labeled with biotin at the 5′ end, were designed and two sets of multiplex asymmetric PCR system were established to amplify the target genes of the seven pathogens. Twelve type specific oligonucleotides were designed and immobilized onto the aldehyde radical modified glass slide to function as target capture probes. After hybridization and stringency washes, the hybridized biotinylated PCR products were detected by the streptavidin-coated magnetic beads. The final hybridization results were visible to the naked eyes and can be imaged by CCD or digital camera. A total of 86 samples previously identified by conventional microbiological methods and/or PCR method were randomly selected to assess the specificity of this assay by a blind study. A coincidence rate of 100% was obtained. Due to the simplicity and specificity of the magnetic bead based DNA microarray, it is especially appropriate for the diagnosis and monitoring of enteric infectious diseases in the community and seaport. © 2010 Springer Science+Business Media B.V. Source

Lu W.,Chongqing Medical University | Gu D.,Research Institute of Disease Control and Prevention | Chen X.,Chongqing Medical University | Xiong R.,Chongqing Medical University | And 3 more authors.
Clinical Chemistry and Laboratory Medicine | Year: 2010

Background: The traditional techniques for diagnosis of invasive fungal infections in the clinical microbiology laboratory need improvement. These techniques are prone to delay results due to their time-consuming process, or result in misidentification of the fungus due to low sensitivity or low specificity. The aim of this study was to develop a method for the rapid detection and identification of fungal pathogens. Methods: The internal transcribed spacer two fragments of fungal ribosomal DNA were amplified using a polymerase chain reaction for all samples. Next, the products were hybridized with the probes immobilized on the surface of a microarray. These species-specific probes were designed to detect nine different clinical pathogenic fungi including Candida albicans, Candida tropocalis, Candida glabrata, Candida parapsilosis, Candida krusei, Candida lusitaniae, Candida guilliermondii, Candida keyfr, and Cryptococcus neoformans. The hybridizing signals were enhanced with gold nanoparticles and silver deposition, and detected using a flatbed scanner or visually. Results: Fifty-nine strains of fungal pathogens, including standard and clinically isolated strains, were correctly identified by this method. The sensitivity of the assay for Candida albicans was 10 cells/mL. Ten cultures from clinical specimens and 12 clinical samples spiked with fungi were also identified correctly. Conclusions: This technique offers a reliable alternative to conventional methods for the detection and identification of fungal pathogens. It has higher efficiency, specificity and sensitivity compared with other methods commonly used in the clinical laboratory. © 2010 by Walter de Gruyter Berlin New York 2010. Source

Gu D.,Research Institute of Disease Control and Prevention | Xie W.,Tsinghua University | Li Z.,PLA Navy Submarine Academy | Lu W.,Chongqing Medical University | And 2 more authors.
Chinese Optics Letters | Year: 2010

Conformations of surface atoms in various stages of nanogold-based genechip testing are scanned by the atomic force and scanning tunneling microscope. We intuitively observe the process and differences in probe combination, nucleic acid hybridization, and silver staining, which might be useful to validate the assay method of genechip. We hope to use this technology to make the other invisible chemical or biochemical reaction become visible and convincible in the future. © 2010 Chinese Optics Letters. Source

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