کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
866937 1470984 2013 7 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
A thermostabilized magnetogenosensing assay for DNA sequence-specific detection and quantification of Vibrio cholerae
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
A thermostabilized magnetogenosensing assay for DNA sequence-specific detection and quantification of Vibrio cholerae
چکیده انگلیسی


► A magnetogenosensor using thermostabilized ready-to-use assay reagents was developed.
► It was successfully used for detection and quantification of pathogenic Vibrio cholerae.
► The thermostabilized assay reagents were stable at room temperature for ≥180 days.
► The magnetogenosensing assay was conducted at room temperature condition.
► The overall assay precision in terms of RSD was <10%, showing its reliability.

Vibrio cholerae is a human pathogen that causes mild to severe diarrheal illnesses and has major public health significance. Herein, we present a thermostabilized electrochemical genosensing assay combining the use of magnetic beads as a biorecognition platform and gold nanoparticles as a hybridization tag for the detection and quantification of V. cholerae lolB gene single-stranded asymmetric PCR amplicons as an alternative to the time-consuming classical isolation method. This thermostabilized, pre-mixed, pre-aliquoted and ready-to-use magnetogenosensing assay simplified the procedures and permitted the reaction to be conducted at room temperature. The asymmetric PCR amplicons were hybridized to a magnetic bead-functionalized capture probe and a fluorescein-labeled detection probe followed by tagging with gold nanoparticles. Electrochemical detection of the chemically dissolved gold nanoparticles was performed using the differential pulse anodic stripping voltammetry method. The real-time stability evaluation of thermostabilized assay was found to be stable for at least 180 days at room temperature (25–30 °C). The analytical specificity of the assay was 100%, while its analytical sensitivity was linearly related to different concentrations of 200-mer synthetic target, purified genomic DNA, and bacterial culture with a limit of detection (LoD) of 3.9 nM, 5 pg/µl, and 103 CFU/ml, respectively. The clinical applicability of the assay was successfully validated using spiked stool samples with an average current signal-to-cut-off ratio of 10.8. Overall, the precision of the assay via relative standard deviation was <10%, demonstrating its reliability and accuracy.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biosensors and Bioelectronics - Volume 47, 15 September 2013, Pages 38–44
نویسندگان
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