Issue link: http://read.dmtmag.com/i/100792
S Be S toc nn el kin u G ec g la tion a Fu ss o ll Bo f ttl es one-seventieth of the product compared to 100%-efficient PCR reaction. In general, assay sensitivity is determined by diluting a target sample in serial dilutions and then testing those dilutions for the presence of the target (e.g. virus). While dilutions up to 10-4 can be detected by well-optimized conventional PCR, qPCR can amplify target from dilutions even up to 10-6 (Figure 2). Our early experiments with qPCR demonstrated this well: When using old RNA samples that had deteriorated during the extended storage in lab freezers, conventional PCR often failed to detect virus in these samples, even though they were previously tested positive by that same method. However, qPCR was still able to show positive signal reliably in these samples. RAISE YOUR same time, ELISA failed to detect virus in 50% of the cases. This difference in detection correlated with the virus concentration: Samples that tested positive by qPCR and negative by ELISA always had Cq values above 25 cycles. DETECTING VIRUS VARIANTS Use of ELISA as a detection method, despite its low sensitivity, has been supported by the argument that it has broad-range detection capability. While this generally applies, it is not always the case. Depending on the type of the antibody (polyclonal or monoclonal) and the source, ELISA reagents��� capability of detecting virus variants can differ significantly (Cohen et al., 2012, Detection of new strains of GLRaV-3 in New Zealand using CHOOSE ALL AMERICAN Find a bottle you love as much as your wine. Make your wine package your very own with a complete line of wine bottles, capsules, screwcaps, labels, shipping supplies, and corks in a variety of colors, shapes, styles, and sizes. Our welcoming sales team can help you with inspiration and direction, answer your questions, and make your purchase easy with fast, clear, and consistent communication. Come see us at booth #1719 at the Unified Wine Symposium Locations in Santa Rosa, CA; San Leandro, CA; Simi Valley, CA; Plano, TX; Belle Vernon, PA; Miami, FL; Tampa, FL; Atlanta, GA; Branchburg, NJ and our newest location in Kalama, WA 78 V I N E YARD & WINERY MANAGEMENT | Figure 2: A series of 10-fold dilutions of GFLV positive RNA sample detected by qPCR. High assay sensitivity is needed when virus titer is low, for example, when detecting viruses in certain rootstock or table grape varieties, or in mixed infections. Enhanced sensitivity is also needed when detecting newly introduced infections of vector-transmitted diseases such as grapevine leafroll or fanleaf (GFLV) virus. Our laboratory���s qPCR results for GFLV from summer 2012 revealed 100,000x differences in the virus titer between the vines in the same vineyard, suggesting active spread of the virus. At the Jan - Feb 2013 ELISA and RT PCR; and Faggioli et al., 2012, Validation of diagnostic protocols for the detection of grapevine viruses covered by phytosanitary rules; proceedings of the 17th Congress of ICVG, Davis Calif.) Well-designed qPCR primers and probes are highly specific and can accurately detect different virus variants without concerns of false negatives or positives. Also, primers designed using the most recent sequence information in the databases can detect more variants than primers published in w w w. v w m m e d i a . c o m