کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
2090717 | 1081515 | 2009 | 5 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: Prion protein detection via direct immuno-quantitative real-time PCR Prion protein detection via direct immuno-quantitative real-time PCR](/preview/png/2090717.png)
We describe a simple and robust assay for the quantitative detection of prions using immuno-quantitative real-time PCR (iQ-RT-PCR) made possible by a direct conjugate of a prion-specific antibody (ICSM35) and a synthetic 99-bp DNA tail. The DNA tail was engineered to include a single ScrFI restriction site, which enabled subsequent quantification of restricted DNA tails using real-time PCR. The assay was tested with scrapie prions bound to polyvinylidene difluoride membranes and to 96-well plates coated with a capturing antibody from a commercially available immuno-based assay (TeSeE). The iQ-RT-PCR assay had a detection limit corresponding to 2.32 × 102 prion epitopes, which represented a 1000-fold increase in detection sensitivity over the commercial assay. Detection of prions from diluted scrapie-positive brain homogenate bound to membranes was linear over a range of 1.06 × 104 to 3.24 × 102 epitopes (R2 = 0.92). Given its sensitivity and versatility, the present assay has potential to enable rapid and reliable detection of agents causing transmissible spongiform encephalopathies.
Journal: Journal of Microbiological Methods - Volume 78, Issue 3, September 2009, Pages 307–311