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Results for “Absolute quantification”

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PubMed1999

Simple high-performance liquid chromatographic method for the determination of tocotrienols in human plasma.

A simple high-performance liquid chromatographic method using fluorescence detection was developed for the determination of vitamin E especially delta-, gamma- and alpha-tocotrienols in human plasma. The method entailed direct injection of plasma sample after deproteinization using a 3:2 mixture of acetonitrile-tetrahydrofuran. The mobile phase comprised 0.5% (v/v) of distilled water in methanol. Analyses were run at a flow-rate of 1.5 ml/min with the detector operating at an excitation wavelength of 296 nm and emission wavelength of 330 nm. This method is specific and sensitive, with a quantification limit of approximately 40, 34 and 16 ng/ml for alpha-, gamma- and delta-tocotrienol, respectively. The mean absolute recovery values were about 98% while the within-day and between-day relative standard deviation and percent error values of the assay method were all less than 12.0% for alpha-, gamma- and delta-tocotrienol. The calibration curve was linear over a concentration range of 40-2500, 30-4000 and 16-1000 ng/ml for alpha-, gamma- and delta-tocotrienol, respectively. Application of the method in a bioavailability study for determination of the above compounds was also demonstrated.

Chromatography, High Pressure Liquid↗

PubMed2005

Calibration of quantitative real-time TaqMan PCR by correlation with hyphal biomass and ITS copies in mycelia of Piloderma croceum.

DNA-based quantification methods such as real-time TaqMan PCR allow a rapid and highly sensitive species-specific quantification of isolated fungal DNA material, but most quantification systems are only able to measure relative amounts of biomass or biomass changes during different treatments. In this experiment, an already established DNA quantification system for the ectomycorrhizal fungus Piloderma croceum, based on the ITS region of ribosomal DNA, was calibrated to absolute biomass to obtain a direct correlation between mycelial biomass and isolated ITS copies. Thin layers of sterile mycelia were cultured on slides. The mycelial biomass was calculated from measurements of the total hyphal length using image analysis, followed by determination of the mycelial volume, and multiplied by the specific weight of hyphae obtained from literature data. Using the very same mycelium, the number of ITS copies was quantified by TaqMan PCR. The mean value of 1047 (+/- 185) copies per mm hypha results in possible data for a direct conversion: one billion (10 (9)) ITS copies corresponded to 0.79 mg hyphal dry weight. For the ribosomal ITS multi-copy genes, the number of ITS copies could be calculated to approx. 152 (+/- 26) copies per dikaryotic cell. These conversion data now allow determination of the mycelial biomass of Piloderma croceum using real-time TaqMan PCR, a prerequisite for competition experiments with Piloderma croceum.

Basidiomycota↗

PubMed1994

Myocardial blood flow at rest and during pharmacological vasodilation in cardiac transplants during and after successful treatment of rejection.

BACKGROUND: The relative intracoronary flow reserve has been found to be reduced during acute transplant rejection, but the effects of rejection on absolute flows at rest and during hyperemia have not been established previously. This has now become possible through noninvasive quantification of myocardial blood flow with positron emission tomography. METHODS AND RESULTS: Myocardial blood flow (MBF) at rest and during dipyridamole-induced hyperemia was quantified in 10 transplant patients (group A) during an acute, biopsy-proven rejection episode and again after successful immunosuppressive treatment and in 6 transplant patients (group B) without prior rejection episode. In group A patients, MBF during rejection averaged 1.7 +/- 0.3 mL.min-1.g-1 at rest and 2.5 +/- 0.9 mL.min-1.g-1 during hyperemia; after recovery, MBF at rest had declined to 1.2 +/- 0.3 mL.-1.g-1 (P < .001) but had increased to 3.9 +/- 1.1 mL.-1.g-1 (P < .001) during hyperemia. Flows after recovery from rejection were similar to those in the group B patients (0.9 +/- 0.2 and 3.9 +/- 0.7 mL.min-1.g-1). Flow reserve in the group A patients was only 1.5 +/- 0.5 during rejection but improved to 3.4 +/- 0.9 at recovery (P < .001) and thus remained lower than in the control patients (4.5 +/- 0.7, P < .05). Minimal coronary resistance during dipyridamole vasodilation was elevated during rejection (40 +/- 11 mm Hg.mL-1.min-1.g-1); after recovery, it no longer differed from that in the group B patients (26 +/- 11 versus 22 +/- 4 mm Hg.mL-1.min-1.g-1). MBF during rejection was increased relative to cardiac work, as demonstrated by significantly higher ratios of blood flow to rate-pressure product than those at recovery and in the control patients. CONCLUSIONS: A decrease in hyperemic and an increase in resting myocardial blood flow, in excess to cardiac work, account for the previously reported reduction in coronary flow reserve. Because both alterations improve with antirejection treatment, they may reflect reversible alterations, presumably of endothelial function, local coagulation, and edema. The compromise in flow reserve and hyperemic flows may contribute to acute and chronic injury from rejection and thus provides a rationale for exercise restriction during rejection. The results further suggest a potential role for serial noninvasive flow measurements to guide immunosuppressive therapy.

Adult↗

PubMed1990

Quantitative radiographic measurement of dentinal lesions.

The purpose of this investigation was the comparison of the precision and accuracy of two reference ramp techniques for the quantification of radiographic density changes in teeth. Radiographs (65 kVp, 10 ma, 1 s, and intra-oral ultraspeed film) of transverse sections from extracted permanent human molars were made before and after dentinal lesions were created. Each radiograph contained the image of a tooth section and the aluminum reference ramp. Method A used the image of the ramp on both the before- and after-lesion radiographs, and method B used the image of the ramp only on the before-lesion radiograph. Three groups of lesions (0.525-mm diameter, n = 11; 0.675-mm diameter, n = 9; and the 0.525-mm holes enlarged to 0.675 mm) were measured radiographically by each technique and by direct planimetry of the lesions. Radiographic method B produced results in close agreement with the planimetric measurements. Method B differentiated (p less than 0.05) between groups that had a mean planimetric size difference of 0.10 mm (equivalent to a change in density difference of 0.6%). These density change measurements are in absolute units of mm of aluminum that can be compared between lesions and between samples. This technique may prove useful for the quantification of changes in mineral density of caries lesions detectable in longitudinal radiographic records.

Dental Caries↗

PubMed2004

A simple mathematical formula for stoichiometry quantification of viral and nanobiological assemblage using slopes of log/log plot curves.

In nanotechnology, biomolecular assemblies serve not only as model systems for the construction of nanodevices, but they can also be used directly as templates for the formation of nanostructures. Biological nano-building blocks can either be isolated as complete functional units from living cells or viruses (biological "Top down" approach) or formed by biomolecular assembly from recombinant or synthetic components ("Bottom up" approach). In both cases, rational design of nanostructures requires knowledge of the stoichiometry of the biological structures, which frequently occur as multimers, i.e., the morphological complex is composed of multiple copies of one or more macromolecules. In this paper, a method is described for the stoichiometric quantification of molecules in bio-nanostructures. The method is based on using dilution factors and relative concentrations rather than absolute quantities, which are often difficult to determine, especially in short-lived assembly intermediates. The approach exploits the fact that the larger the stoichiometry of the component is, the more dramatic is the influence of the dilution factor (decrease in concentration) on the reaction. We established and used the method to determine the stoichiometry of components of bacterial virus phi29. The log of dilution factors was plotted against the log of reaction yield. The stoichiometry Z was determined with the equation Z=-1.58+2.4193T-0.001746T(2) [T in (0,1000), or 90 degree angle alpha in (0 degrees, 89.9 degrees )], where T is the slope of the curve (tangent of 90 degree angle alpha, which is the angle between the x-axis and the concentration dependent curve). Z can also be determined from a standard table given in this report. With the bacteriophage phi29 in vitro assembly system, up to 5x10(8) infectious virions per ml can be assembled from 11 purified components, giving our method a sensitivity of nine orders of magnitude. We confirmed the stoichiometries of phi29 components that were determined previously with microscopic approaches. The described method also responded to programmed stoichiometry changes, which were generated by assembling the phi29 DNA packaging motor from modified pRNA (DNA-packaging RNA) molecules forming a trimer of dimers or a dimer of trimers, instead of the wild-type hexamer.

Bacillus Phages↗

PubMed2000

Vessel diameter measurements in gadolinium contrast-enhanced three-dimensional MRA of peripheral arteries.

In this study, the possibilities for quantification of vessel diameters of peripheral arteries in gadolinium contrast-enhanced magnetic resonance angiography (Gd CE MRA) were evaluated. Absolute vessel diameter measurements were assessed objectively and semi-automatically in maximum intensity projections (MIPs) of contrast-enhanced T1-weighted 3D spoiled gradient-echo datasets, studied with digital subtraction techniques. In vivo, the complete peripheral arterial bed of six patients was studied, from the aorto-iliac bifurcation down to the distal run-off. By measuring the signal intensity (SI) over the lumen of a vessel in the MIP, an SI-plot was obtained. Next, the vessel boundaries were determined using a threshold algorithm; from these boundary points individual diameter values could be obtained along the trajectory of the vessel. In an in vitro study, an optimal threshold value of 30% of the range of SI-values between the background and the maximal SI in the vessel was obtained for accurate diameter measurement in Gd CE MRA (i.e., full-width 30%-maximum). Furthermore, the relationship between the accuracy of these measurements and the scan resolution was investigated. Accuracy was found to be acceptable (i.e., less than 10% over/underestimation) for vessel sizes covering at least 3 pixels. In six patients, diameters were measured in MIPs of the total datasets (i.e., D(T)) as well as in selective MIPs of the clipped datasets (i.e., D(S)) (n = 209). D(T) and D(S) were statistically significantly correlated (p < 0.01) with a Pearson correlation coefficient rP = 0.98. Measurements in the total MIPs yielded statistically significant (p < 0.01) smaller diameter values compared with measurements in selective MIPs, with a mean difference of 0.15 mm. Diameter values from the selective MIPs of the aorto-iliac arteries were also compared with diameter values measured at corresponding anatomic positions in X-ray angiograms of these patients (i.e., D(x)) (n = 70). D(X) and D(S) were statistically significantly correlated (p < 0.01) with a Pearson correlation coefficient rP = 0.92. Diameters measured in the selective MIPs were smaller than those measured in the X-ray angiograms (mean difference 0.49 mm) and this difference was statistically significant (p < 0.01). In conclusion, diameter values can be evaluated accurately in MIPs of vessels with at least 3 pixels in diameter, using the full-width 30%-maximum criterion.

Adult↗

PubMed2004

Identification and quantification of gossypol in cotton by using packed micro-tips columns in combination with HPLC.

Self-packed micro-tip columns containing a C18-bonded silica stationary phase, based on the same principles as solid-phase extraction methods, were used to obtain gossypol and related sesquiterpenoid aldehyde-enriched fractions. The enriched metabolite fractions were then analyzed by optimized high-performance liquid chromatography (HPLC) with a C18 column (4.6 mm x 25 cm) eluted with the binary mobile phase acetonitrile-0.1% aqueous TFA solution (80:20). This method has proven to be highly reproducible. The precision and accuracy, as %RSD and %RME values, were determined to be less than 15% for the method. The minimum detection limit of gossypol was determined to be 10 ng (absolute gossypol). Absolute recovery was greater than 94% with a standard deviation of +/-3.68%. This is a simple, fast, and cost-effective method for isolation, identification, and quantification of gossypol and related secondary metabolites. Comparative analysis of gossypol content was performed on different parts of the cotton plant (seeds, stems and leaves) of two different cultivars of Gossypium hirsutum L. (Acala(1517-70) and OR19). The results indicate that the OR19 cv naturally contains higher gossypol levels than the Acala cv. It was also found that treatment of leaves with a Verticillium dahliae-derived elicitor induced production of deoxyhemigossypol rather than gossypol.

Chromatography, High Pressure Liquid↗

PubMed1998

Design and validation of a new image analysis method for automatic quantification of interstitial fibrosis and glomerular morphometry.

Interstitial fibrosis and morphologic changes in kidney glomeruli, the structural effects of many diseases, lead to significant pathologic alterations. A reliable and objective method to accurately quantify the extent of interstitial fibrosis and the degree of alteration in glomerular morphology is needed for both clinical practice and experimental work. The morphometric methods of quantification described to date are time-consuming and require trained personnel. This article describes the design and validation of an image analysis-based application (Fibrosis HR) for automatically and rapidly quantifying interstitial fibrosis and glomerular morphology in the same tissue section stained with Sirius red. The image processing algorithms described herein automatically segment interstitial fibrosis and mesangial matrix using automatic thresholding and morphologic filtering. The glomerular region is extracted by a simple interactive step and an automatic mathematical morphology algorithm, whereas the glomerular tuft is automatically segmented with automatic thresholding and a sequence of Boolean and mathematical morphology operations. All extracted areas are automatically quantified in absolute (microm2) and relative (%) values. For validation of this method, interstitial fibrosis, mesangial matrix, and glomerular and glomerular tuft areas were manually segmented and their quantifications statistically compared with those obtained automatically. Statistical analyses showed significant intra- and interoperator variability in manual segmentation of interstitial fibrosis, mesangial matrix, and glomerular tuft areas. Automatic quantifications of the same areas did not differ significantly from their mean manual evaluations. There was no significant intra- or interoperator variability in the interactive identification of the glomerular region. In conclusion, Fibrosis HR produces robust, fully reproducible, accurate, objective, and reliable quantifications, which facilitate the evaluation of in vivo experimental models of renal interstitial and glomerular pathologies and improve the accuracy of clinicopathologic analyses of renal diseases in human biopsies.

Algorithms↗

PubMed1997

Comparison of competitive and non-competitive reverse transcription-polymerase chain reaction (RT-PCR) for the quantification of hepatitis C virus (HCV) RNA.

A competitive reverse transcription (RT)-nested polymerase chain reaction (PCR) assay for HCV RNA was compared with the Roche Amplicor HCV Monitor assay, based on non-competitive, single step RT-PCR. A total of 83 serum samples were tested in parallel by both assays. All samples could be quantified by competitive RT-PCR (cPCR), whereas seven were negative by the non-competitive assay (ncPCR). The HCV RNA titre of the discordant samples assessed by cPCR was significantly lower than that of the remaining 76 (P < 0.001). Absolute HCV RNA titres were higher by cPCR than by ncPCR (P < 0.001), even if the results of the two methods were statistically correlated (P < 0.001). HCV RNA titre tested by cPCR was not significantly different between samples infected with genotype 1 or 2. However, values obtained by ncPCR were higher in samples with genotype 1 (P < 0.001). Furthermore, all seven discordant samples were infected with genotype 2. When both methods were used to measure serial dilutions of standard HCV RNA, we observed a bias to lower measurements with the ncPCR kit. This study shows a good correlation between the results of two PCR-based methods for the quantification HCV RNA. However, the degree of sensitivity and the absolute HCV RNA titre measured may vary according to the assay used.

Analysis of Variance↗

PubMedDate not supplied

Quantification of SPECT myocardial perfusion images: methodology and validation of the Yale-CQ method.

BACKGROUND: Quantification of single photon emission computed tomography (SPECT) images is important for reproducible and accurate image interpretation. In addition, SPECT quantification provides important prognostic information. The purpose of this study was to validate the Yale circumferential quantification (Yale-CQ) method in phantom studies. METHODS: Myocardial perfusion defects of varying extent and severities were simulated in a cardiac phantom with fillable defect inserts. Forty-five different phantom configurations simulated 45 different myocardial perfusion defect sizes, ranging from 1.6% to 32% of the cardiac phantom volume. Automatic processing was compared with manual processing in the phantom SPECT studies. RESULTS: The automatic Yale-CQ algorithm performed well in all phantom studies. Compared with manual processing, the mean absolute error for automatically determined center of short axis slices was 0.27 pixel in the x direction, 0.45 pixel in the y direction, and 0.15 pixel in radius. Quantification of phantom defects with the Yale-CQ method correlated well with actual defect sizes (R = 0.99), but there was a systematic underestimation (mean error = -7.9%). With derived correction factors the overall correlation between 45 phantom defects and actual defect sizes was excellent, and the estimation error was significantly improved (R = 0.98, mean error = -0.82% for manual method and -0.95% for automatic method). CONCLUSION: The automatic processing algorithm performs well for the phantom studies. Myocardial perfusion abnormalities can be quantified accurately by use of the Yale-CQ method. Quantified SPECT defect size can be expressed as a percentage of the left ventricle.

Algorithms↗

PubMed1995

High-performance liquid chromatographic determination of urinary 4'-hydroxymephenytoin, a metabolic marker for the hepatic enzyme CYP2C19, in humans.

The preferential hydroxylation of (S)-mephenytoin to 4'-hydroxymephenytoin (4'-OH-M) displays a genetic polymorphism of drug metabolism in humans. Thus the excreted 4'-OH-M is considered to be an important marker for the hepatic (S)-mephenytoin 4'-hydroxylase. Accordingly, a mixture of urine containing total 4'-OH-M after enzymatic deconjugation and phenobarbital as internal standard (I.S.) was extracted with absolute diethyl ether. The residue remaining after evaporation was dissolved in 50 microliters of eluate and 20 microliters were injected into the chromatographic system. All components were separated isocratically on a reversed-phase column using acetonitrile-water (24:76, v/v) as the mobile phase at a flow-rate of 1.2 ml/min. The effluent was monitored at 204 nm. The retention times for 4'-OH-M and the I.S. were within 6 min. The absolute recovery was in the range 84-89% for 4'-OH-M and that of the I.S. was 75.9 +/- 4.2%. Quantification was performed by measuring the peak-height ratio compared with the ratio of the amount of 4'-OH-M divided by that of the I.S. The intra- and inter-day variations were less than 8% and 10%, respectively. The proposed method is simpler and more convenient than those reported previously. Its practical applicability was assessed by phenotyping the efficient and deficient hydroxylators among the Chinese minorities and Han Chinese populations.

Aryl Hydrocarbon Hydroxylases↗

PubMed1996

Quantification of dextromethorphan and metabolites: a dual phenotypic marker for cytochrome P450 3A4/5 and 2D6 activity.

A sensitive and selective liquid chromatographic procedure using fluorimetric detection was developed to quantify dextromethorphan (DTM), 3-methoxymorphinan (3MM), dextrorphan (DT), 3-hydroxymorphinan (3OH) and two internal standards, codeine (COD) and ethylmorphine (ETM), in urine. Precision and accuracy of the assay were determined over a concentration range of 5-3200 ng/ml urine for DTM, 5-400 ng/ml urine for 3MM, 400-40 000 ng/ml urine for DT and 200-16 000 ng/ml urine for 3OH, by assaying freshly prepared calibration standards and replicates of six quality control (QC) samples on separate days. All of the inter-day and intra-day coefficients of variation (C.V.s) were less than 20% except for a low QC for 3MM. The inter-day and intra-day accuracies were less than 20% for the low QCs, less than 15% for the medium QCs and less than 12% for the high QCs, for all compounds. The limit of quantification (LOQ) was 2 ng/ml urine for DTM and 3MM, 250 ng/ml urine for DT, and 100 ng/ml urine for 3OH. Absolute recovery was 76% for DTM, 74% for 3MM, 77% for DT, 46% for 3OH, 73% for ETM, and 57% for COD. The frequency distribution of the CYP2D6 metabolic ratio (DTM/DT) illustrated a bimodal distribution whereas, the CYP3A metabolic ratio (DTM/3MM) exhibited a unimodal distribution in overnight urine samples of volunteers who ingested 30 mg dextromethorphan hydrobromide. The CYP2D6 metabolic ratio significantly correlated with 3MM/3OH (r=0.82) and DTM/3OH (r=0.95) but did not correlate with the CYP3A metabolic ratio (r=0.27).

Chromatography, High Pressure Liquid↗

PubMed2006

A HPLC-mass spectrometric method suitable for the therapeutic drug monitoring of everolimus.

We report here the validation of an HPLC-electrospray-tandem mass spectrometry method for the quantification of everolimus, an immunosuppressant drug. Whole blood samples (100 microl) were extracted by protein precipitation which involved sample pre-treatment with zinc sulphate followed by acetonitrile (containing internal standard, 40-O-(3'-hydroxy)propyl-rapamycin). HPLC was performed using a step-gradient at a flow rate of 0.6 ml/min on a Waters TDM C18 column (10 mm x 2.1mm I.D.) with a resultant chromatographic analysis time of 2 min. Mass spectrometric detection by selected reaction monitoring (everolimus m/z 975.5-->908.3; internal standard m/z 989.5-->922.3). The assay was linear from 0.5 to 40 microg/l (r2>0.994, n=11). The inter- and intra-day analytical recovery and imprecision for quality control samples (1.25, 12.5 and 30 microg/l) were 93.4-98.2% and <10.7%, respectively (n=10). At the lower limit of quantification (0.5 microg/l) the inter- and intra-day analytical recovery was 94.4-95.8% with imprecision of <14.1% (n=10). The absolute recovery of everolimus (6.5 microg/l) and internal standard (12.5 microg/l) was 96.5 and 88.3%, respectively (n=3). A comparison of our method against the mean of all HPLC methods for a series of samples from an external proficiency testing scheme revealed good correlation as shown by the regression analysis: y=0.973x+0.301 (r2=0.986, n=71). In conclusion, the method described is suited to the current requirements for therapeutic drug monitoring of everolimus.

Chromatography, High Pressure Liquid↗

PubMed2005

Development and validation of a bioanalytical method using automated solid-phase extraction and LC-UV for the simultaneous determination of lumefantrine and its desbutyl metabolite in plasma.

A bioanalytical method for the determination of lumefantrine (LF) and its metabolite desbutyl-lumefantrine (DLF) in plasma by solid-phase extraction (SPE) and liquid chromatography has been developed. Plasma proteins were precipitated with acetonitrile:acetic acid (99:1, v/v) containing a DLF analogue internal standard before being loaded onto a octylsilica (3 M Empore) SPE column. Two different DLF analogues were evaluated as internal standards. The compounds were analysed by liquid chromatography UV detection on a SB-CN (250 mm x 4.6 mm) column with a mobile phase containing acetonitrile-sodium phosphate buffer pH (2.0; 0.1 M) (55:45, v/v) and sodium perchlorate 0.05 M. Different SPE columns were evaluated during method development to optimise reproducibility and recovery for LF, DLF and the two different DLF analogues. The within-day precisions for LF were 6.6 and 2.1% at 0.042 and 8.02 microg/mL, respectively, and for DLF 4.5 and 1.5% at 0.039 and 0.777 microg/mL, respectively. The between-day precisions for LF were 12.0 and 2.9% at 0.042 and 8.02 microg/mL, respectively, while for DLF 0.7 and 1.2% at 0.039 and 0.777 microg/mL, respectively. The limit of quantification was 0.024 and 0.021 microg/mL for LF and DLF, respectively. Different amounts of lipids in plasma did not affect the absolute recovery of LF or DLF.

Chromatography, Liquid↗

PubMed1994

Quantification of CD24 and CD45 antigens in parallel allows a precise determination of B-cell maturation stages: relevance for the study of B-cell neoplasias.

Quantitative expression, i.e. absolute number of monoclonal antibody molecules bound per cell, was evaluated for CD24 and CD45 by flow cytometry with standards of fluorescence intensity on a panel of normal and neoplastic B cells. The CD24 antigen was expressed at homogeneous high level in fetal bone marrow and liver. Its density decreased progressively in the other normal tissues in parallel with the B-cell maturation. The ratio between CD24 density measured on fetal bone marrow B cells and that seen on adult peripheral B cells was 6:1. The CD45 antigen density was lower on fetal bone marrow cells than in the more mature stages. Fetal spleen lymphocytes and all the mature B lymphocytes displayed the same CD45 density than that seen on normal adult peripheral T cells. The CD24/CD45 antigen density ratio was precisely related to the stage of B-cell maturation. The same pattern of variation of CD24 and CD45 antigen density was seen on B-cell neoplasias, with a significantly higher value of CD24 and lower value of CD45 in acute lymphoblastic leukemias than in chronic malignancies. CD24 and CD45 antigen levels were frequently out of the range observed in the corresponding normal population. Among ALL patients, a low CD24/CD45 antigen density ratio was associated with a good prognosis. These data confirm the interest of an absolute quantitative study for widely distributed antigens.

Antigens, CD↗

PubMed2002

Echocardiographic quantification of left ventricular asynchrony predicts an acute hemodynamic benefit of cardiac resynchronization therapy.

OBJECTIVES: We sought to determine whether radial left ventricular (LV) asynchrony in patients with heart failure predicts systolic function improvement with cardiac resynchronization therapy (CRT). BACKGROUND: We quantified LV wall motion by echocardiography to correlate the effects of CRT on LV systolic function with wall motion synchrony. METHODS: Thirty-four patients underwent echocardiographic phase analysis of LV septal and lateral wall motion and hemodynamic testing before CRT. Phase relationships were measured by the difference between the lateral (Phi(L)) and septal (Phi(S)) wall motion phase angles: Phi(LS) = Phi(L) - Phi(S). The absolute value of Phi(LS) was used as an order-independent measure of synchrony: the absolute value Phi(LS) = the absolute value of Phi(L) - Phi(S). RESULTS: Three phase relationships were identified (mean +/- SD): type 1 (n = 4; peak positive LV pressure [dP/dt(max)] 692 +/- 310 mm Hg/s; Phi(LS) = 5 +/- 6 degrees, synchronous wall motion); type 2 (n = 17; dP/dt(max) 532 +/- 148 mm Hg/s; Phi(LS) = 77 +/- 33 degrees, delayed lateral wall motion); and type 3 (n = 13; dP/dt(max) 558 +/- 154 mm Hg/s; Phi(LS) = -115 +/- 33 degrees, delayed septal wall motion, triphasic). A large absolute value of Phi(LS) predicted a larger increase in dP/dt(max) with CRT (r = 0.74, p < 0.001). Sixteen patients were studied during right ventricular (RV), LV and biventricular (BV) pacing. Cardiac resynchronization therapy acutely reduced the absolute value of Phi(LS) from 104 +/- 41 degrees (OFF) to 86 +/- 45 degrees (RV; p = 0.14 vs. OFF), 71 +/- 50 degrees (LV; p = 0.001 vs. OFF) and 66 +/- 42 degrees (BV; p = 0.001 vs. OFF). A reduction in the absolute value of Phi(LS) predicted an improvement in dP/dt(max) in type 2 patients for LV (r = 0.87, p = 0.005) and BV CRT (r = 0.73, p = 0.04). CONCLUSIONS: Echocardiographic quantification of LV asynchrony identifies patients likely to have improved systolic function with CRT. Improved synchrony is directly related to improved hemodynamic systolic function in type 2 patients.

Aged↗

PubMed2003

Validation of an algorithm for automatic quantification of nucleic acid copy numbers by real-time polymerase chain reaction.

Real-time quantitative polymerase chain reaction (PCR) with on-line fluorescence detection has become an important technique not only for determination of the absolute or relative copy number of nucleic acids but also for mutation detection, which is usually done by measuring melting curves. Optimum assay conditions have been established for a variety of targets and experimental setups, but only limited attention has been directed to data evaluation and validation of the results. In this work, algorithms for the processing of real-time PCR data are evaluated for several target sequences (p53, IGF-1, PAI-1, Factor VIIc) and compared to the results obtained by standard procedures. The algorithms are implemented in software called SoFAR, which allows fully automatic analysis of real-time PCR data obtained with a Roche LightCycler instrument. The software yields results with considerably increased precision and accuracy of quantifications. This is achieved mainly by the correction of amplification-independent signal trends and a robust fit of the exponential phase of the signal curves. The melting curve data are corrected for signal changes not due to the melting process and are smoothed by fitting cubic splines. Therefore, sensitivity, resolution, and accuracy of melting curve analyses are improved.

Algorithms↗

PubMed1999

Differentiation of small hyperechoic renal cell carcinoma from angiomyolipoma: computer-aided tissue echo quantification.

To assess the value of computer-aided tissue echo quantification in differentiating small hyperechoic renal cell carcinomas from angiomyolipomas, we studied ultrasonographic images of 15 renal cell carcinomas and 20 angiomyolipomas. After digitizing the images, we measured the absolute gray scale values of the renal cortex, central echo complex, and mass. The relative gray scale value (%) of the mass was calculated by setting the gray scale value of the cortex as 0% and the central echo complex as 100%. The relative gray scale value of renal cell carcinomas was in the range of 12 to 73% (mean, 28%), whereas that of angiomyolipomas was 30 to 204% (mean, 130%). The differentiation between small hyperechoic renal cell carcinomas and angiomyolipomas can be facilitated by computer-aided tissue echo quantification.

Adult↗