کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10532963 | 961807 | 2010 | 6 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Enhanced detection of staphylococcal genomes in positive blood cultures using a polymeric enzyme complex
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
This article describes a simple and inexpensive signal amplification method, termed polymeric enzyme detection (PED), which permits rapid and sensitive detection of conserved sequences in the tuf gene that identify Staphylococcus genus, conserved sequences in the femB gene that specifically detect Staphylococcus aureus species, and the methicillin resistance gene mecA directly from positive blood culture bottles. Microbe-specific capture probes were immobilized onto microtiter plates or silicon chips. Target sequences and biotin-labeled, target-specific probes were hybridized to complementary capture probes to create a biotin-labeled, surface-immobilized tripartite complex. In a two-step process, signal was amplified by incubating the surface-immobilized biotin with streptavidin followed by the addition of a 500-kDa dextran polymer conjugated with approximately 80 biotins. Signal was then developed by binding of a streptavidin-horseradish peroxidase conjugate followed by incubation with the substrate tetramethylbenzidine. Use of the PED method improved the lower limit of detection 10- to 100-fold in model DNA hybridization assays with limits of detection as low as 1Â fmol/L target DNA. This level of sensitivity permits detection of genomic DNA from methicillin-resistant S. aureus positive blood cultures within 25 to 35Â min using either a thin film biosensor chip or a microtiter plate-based assay.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Analytical Biochemistry - Volume 396, Issue 2, 15 January 2010, Pages 284-289
Journal: Analytical Biochemistry - Volume 396, Issue 2, 15 January 2010, Pages 284-289
نویسندگان
Joshua Klonoski, Roy Mondesire, Larry Rea, David C. Ward, Robert D. Jenison,