1 RealTime ready Assay Production and Function Testing 2 Production 2 Function Testing 2 A Census on 10,000+ RealTime ready Assays 6 Selecting the Design Sequence 6 Assay Design 6 Amplification Efficiencies 8 Bibliography References 9 M. Dietrich, U. H. Grepl, B. Kramer and H. Walch Roche Diagnostics GmbH Nonnenwald 2 82377 Penzberg / Germany RealTime ready RT-qPCR Assay Development and Qualifi...

Epigenetic markers based on differential methylation of DNA sequences are used in cancer screening and diagnostics. Detection of abnormal methylation at specific loci by real-time quantitative polymerase chain reaction (RT-qPCR) has been developed to enable high-throughput cancer screening. For tests that combine the results of multiple PCR replicates into a single reportable result, both indiv...

Crimean-Congo hemorrhagic fever (CCHF) is a zoonotic disease caused by a nairovirus belonging to family Bunyaviridae. The CCHF virus (CCHFV) can be transmitted to humans by Hyalomma ticks as well as by direct contact with infected body fluids or tissues from viremic livestock or humans. Our aim was to set up a fast RT-qPCR for detection of the different CCHFV genotypes in clinical samples, incl...

This Application Note describes and demonstrates the efficiency of the Agilent 2200 TapeStation system and the RNA ScreenTape assay in RT-qPCR workflows for assessing the integrity of the initial total RNA template. This template is crucial for the successful amplification of the target genes. Moreover, the D1000 ScreenTape assay helps assess the amplified products for sizing and eliminate fals...

Ongoing evolution of viral pathogens is a significant issue in diagnostic virology employing TaqMan qPCR/RT-qPCR. Specific concerns are related to false negativity due to probe binding failure. One option for compensating for such deficiency is to integrate a second identically labelled probe in the assay. However, how this alteration influences the reaction parameters has not been comprehensiv...

Real-time PCR, or quantitative PCR (QPCR), has been developed to rapidly detect and quantify the total number of yeasts in wine without culturing. Universal yeast primers were designed from the variable D1/D2 domains of the 26S rRNA gene. These primers showed good specificity with all the wine yeasts tested, and they did not amplify the most representative wine species of acetic acid bacteria a...

Individual cells represent the basic unit in tissues and organisms and are in many aspects unique in their properties. The introduction of new and sensitive techniques to study single-cells opens up new avenues to understand fundamental biological processes. Well established statistical tools and recommendations exist for gene expression data based on traditional cell population measurements. H...

Excessive subconjunctival scarring is the main reason of failure of glaucoma filtration surgery. We analyzed conjunctival and systemic gene expression patterns after non penetrating deep sclerectomy (NPDS). To find expression patterns related to surgical failure and their correlation with the clinical outcomes. This study consisted of two consecutive stages. The first was a prospective analysis...

Sensitive detection of water- and foodborne enteric viruses is extremely relevant, especially due to the low concentrations in which they are found. Accurate and sensitive detection of Norovirus, the primary responsible for water- and foodborne outbreaks, is of particular importance. Quantification of Norovirus is commonly performed by quantitative RT-PCR (RT-qPCR). In recent years a new platfo...

HIGHLIGHTS RT-qPCR efficiency can vary according to 260/280 ratio of samples analyzed. Chemotherapeutic drugs in the blood sample was not correlated with Ct variation (qPCR). The spectrophotometer determines a RNA quantification 2.2 times higher than fluorimeter. Antibiotics be associated 11.3% variability ratio.