In this multicenter evaluation, the VERSANT HCV RNA 3.0 Assay (bDNA) (Bayer Diagnostics, Tarrytown, N.Y.) was shown to have excellent reproducibility, linearity, and analytical sensitivity across specimen collection matrices (serum, EDTA, ACD-A), and hepatitis C virus (HCV) genotypes 1 to 6. The VERSANT HCV bDNA Assay has a reportable range of 615 to 7,690,000 (7.69 ؋ 10 6 ) IU/ml. The total coefficient of variation (CV) ranged from 32.4% at 615 IU/ml to 17% at 6.8 ؋ 10 6 IU/ml. The assay was linear across the reportable range. Analytical specificity of 98.8% was determined by testing 999 specimens from volunteer blood donors. Evaluation of HCV genotypes using RNA transcripts of representative clones of 1a, 1b, 2a, 2b, 2c, 3a, 4a, 5a, and 6a and patient specimens showed that the largest difference between genotype 1, upon which the assay is standardized, and non-1 genotypes was within 1.5-fold. Testing of potentially interfering endogenous substances and exogenous substances and conditions found no interference in HCV-positive or HCV-negative specimens except for unconjugated bilirubin at concentrations of >20 mg/dl and protein at concentrations of >9 g/dl. Biological variability was estimated from 29 clinically stable individuals not on HCV therapy who were tested weekly over an 8-week period. The combined estimate of total (biologic plus assay) variability was 0.15 log 10 standard deviation (CV, 36.1%), a fold change of 2.6. Thus, the observed fold change between any two consecutive HCV RNA measures is expected to be less than 2.6-fold (equivalent to 0.41 log 10 IU/ml) 95% of the time in clinically stable individuals.Hepatitis C virus (HCV) infection is a major health care problem, with an estimated 4 million individuals infected in the United States (1) and 170 million infected worldwide (26). Approximately 70% of those infected will develop chronic HCV infection, which is a leading cause of chronic liver disease (13). Interferon monotherapy was the first antiviral regimen approved for treatment of chronic HCV infection. Sustained response rates were low (8 to 12%) (3), but newer antiviral treatments (interferon plus ribavirin, peginterferon plus ribavirin) have improved the response rates, particularly for patients infected with non-1 genotypes.Quantitative HCV RNA results have been used prior to initiating interferon-based anti-HCV treatment to assess the likelihood of sustained virological response, defined as a negative result in a qualitative HCV RNA assay 6 months after the end of treatment. HCV RNA levels or changes from baseline have also been used early in treatment to attempt to accurately predict response to treatment (6,10,14,15,18,20,27).The performance characteristics of an assay, i.e., its accuracy, reproducibility, and predictive values, determine the interpretation and utility of the assay's results (12, 18). The VERSANT HCV bDNA Assay, utilizing a signal amplification technique, is a fundamentally simple hybridization-based procedure involving unique nucleic acid probes directed to highly co...
Infection with hepatitis B virus (HBV) remains a difficult worldwide challenge to public health. The World Health Organization estimates that more than one-third of the world's population has been infected with HBV (25). Epidemiological trends suggest there are currently 400 million HBV chronic carriers worldwide, with over 1 million deaths annually due to HBV-associated liver disease (13,23). HBV is the leading cause of cirrhosis and hepatocellular carcinoma globally (25; Centers for Disease Control and Prevention hepatitis fact sheet [www.cdc.gov/hepatitis]). The development and utilization of molecular diagnostic assays for the detection and quantification of HBV genomes have provided insight into the natural history of HBV and the pathogenesis of HBV infection as well as facilitated the monitoring of viral response to treatment (15,17). In addition, a quantitative evaluation of HBV DNA concentrations can provide valuable information on the levels of viral replication and may be useful as a prognostic indicator of liver disease (4, 21). A number of commercial assays are currently available for the quantification of HBV DNA in patient serum or EDTA-plasma, including hybridization-, signal-, and target-amplification-based technologies (5,11,(16)(17)(18)22). Selection of the optimal assay is dependent on the intrinsic performance characteristics of the methodology as well as the necessity to make appropriate clinical decisions in the context of HBV-associated disease (4, 15, 21).The VERSANT HBV 3.0 Assay (referred to herein as VER-SANT 3.0) is a third-generation branched-DNA (bDNA) assay for the direct quantification of HBV DNA in human serum and plasma. After HBV genomic DNA is released from the virions, the viral DNA is captured by a set of specific, synthetic oligonucleotide capture probes fixed in a microtiter well. A set of target probes (or label extender probes) then hybridizes to both the captured viral DNA and unique preamplifier probes. The capture probes and the target probes bind to conserved DNA regions throughout the entire HBV genome. The amplifier probes subsequently hybridize to the preamplifier probes, forming a bDNA complex. Multiple copies of an alkaline phosphatase-labeled probe are then hybridized to this immobilized complex. Detection is achieved by incubating the alkaline phosphatase-bound complex with a chemiluminescent substrate. The intensity of light emission is directly related to the amount of HBV DNA present in each sample, and results are recorded as relative light units by the luminometer. A standard curve is defined by light emission from quantitative standards containing known concentrations of recombinant DNA. Concentrations of HBV DNA in specimens are determined from this standard curve. This third-generation sandwich nucleic acid hybridization procedure differs from earlier bDNA assays by using the unique preamplifier probes to increase the number of labeled probes that can bind to the target, thereby
Branched DNA (bDNA) is a signal amplification technology used in clinical and research laboratories to quantitatively detect nucleic acids. An overnight incubation is a significant drawback of highly sensitive bDNA assays. The VERSANT® HIV-1 RNA 3.0 Assay (bDNA) (“Versant Assay”) currently used in clinical laboratories was modified to allow shorter target incubation, enabling the viral load assay to be run in a single day. To dramatically reduce the target incubation from 16–18 h to 2.5 h, composition of only the “Lysis Diluent” solution was modified. Nucleic acid probes in the assay were unchanged. Performance of the modified assay (assay in development; not commercially available) was evaluated and compared to the Versant Assay. Dilution series replicates (>950 results) were used to demonstrate that analytical sensitivity, linearity, accuracy, and precision for the shorter modified assay are comparable to the Versant Assay. HIV RNA-positive clinical specimens (n = 135) showed no significant difference in quantification between the modified assay and the Versant Assay. Equivalent relative quantification of samples of eight genotypes was demonstrated for the two assays. Elevated levels of several potentially interfering endogenous substances had no effect on quantification or specificity of the modified assay. The modified assay with drastically improved turnaround time demonstrates the viability of signal-amplifying technology, such as bDNA, as an alternative to the PCR-based assays dominating viral load monitoring in clinical laboratories. Highly sensitive bDNA assays with a single day turnaround may be ideal for laboratories with especially stringent cost, contamination, or reliability requirements.
The multivariate extension of control charts for process dispersion is not as straightforward as that for the process mean. A general model and techniques that would encompass a wide range of problems encountered in practice is not available. In most cases, particular problems need to be handled in a specific manner. In this paper we consider several special cases of a process displacement affecting the covariance matrix and we develop control charts (both Shewharttype and CUSUM) to detect these process changes.
Economic design of control charts seeks to choose the parameters of control charts that will minimizethe cost. In an economic model, the time to the occurence of an assignable cause is usually assumed to follow an exponential distribution. In addition to this assumption, certain approximations are used in the optimization procedures. This paper extends the current economic design for X charts in two directions. First, economic design is considered for a general distribution for the time to the occurrence of an assignable cause. The method is specialized to several distributions such as Weibull, lognormal, folded-normal, folded-logistic, and gamma. Second, an exact method to obtain the optimal design parameters is developed. Using Duncan's examples, the optimal designs obtained by exact method are compared for different distributions. These are also compared with the approximate optimal designs given by Duncan. This comparison shows that the optimality of the design for X charts is insensitive to the distributional assumption ofthe time to the occurrence of an assignablecause for small valuesof the sampling interval.
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