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The "X-Ray RheumaCoach" software: a novel tool for enhancing the efficacy and accelerating radiological quantification in rheumatoid arthritis.

BACKGROUND: Precise diagnosis and follow up treatment of rheumatoid arthritis (RA) requires objective quantification, which is still lacking. For this purpose, radiological analyses are considered to be the most appropriate method. OBJECTIVE: To develop computer assisted quantification software that is particularly applicable to joint scoring in rheumatic disorders. METHODS: 3914 radiographs from hands and feet of 190 patients with RA were collected, expertly examined, analysed, and statistically evaluated. Radiographs were quantified using the conventional Larsen score and the "X-Ray RheumaCoach" (XRRC) software. The XRRC is a Java stand alone application which can support and accelerate, but not fully automate, the scoring procedure in RA. The scorer can apply both the Larsen and the Ratingen-Rau scores. RESULTS: Compared with conventional scoring procedures, the XRRC software accelerated quantification time by approximately 25%. The program, which is now available on the internet free of charge, ran stably and proved to be a consistently valuable tool. CONCLUSIONS: Compared with conventional scoring methods, the XRRC software offers several advantages: (a) structured data analysis and input that minimises variance by standardisation; (b) faster and more precise calculation of sum scores and indices; (c) permanent data storing and fast access to the software's database; (d) the possibility of cross calculation to other scores; (e) "user friendly" technology and a dedicated help program; (f) fast access and data transfer through the internet if desired; and (g) reliable documentation of results in a specially designed printout.

Arthritis, Rheumatoid↗

Coronary artery calcium: absolute quantification in nonenhanced and contrast-enhanced multi-detector row CT studies.

PURPOSE: (a) To determine the accuracy of multi-detector row computed tomography (CT) in the measurement of the calcium concentration in a cardiac CT calibration phantom and (b) to assess the correlation of a traditional 3-mm section width CT coronary screening protocol and a 1.25-mm section width CT angiography imaging protocol in the quantification of the absolute mass of coronary calcium in patients who underwent both coronary screening and CT angiography with a multi-detector row CT scanner. MATERIALS AND METHODS: A heart phantom containing calcified cylinders was scanned to determine calibration factors and absolute calcium mass. In 50 patients, the variability (value 1 - value 2/mean value 1 - value 2), limit of agreement (+/-2SD value 1 - value 2), and systematic error (mean value 1 - value 2) of the total amount of coronary calcium calculated at traditional 3-mm section width CT and at 1.25-mm section width CT angiography were determined. RESULTS: The correlation coefficient between the 3-mm section width, nonenhanced protocol and the 1.25-mm section width CT angiography protocol was very high (r = 0.977) and the mean variability was low (19.7%) for the absolute mass. There was a systematic error of -6.7 mg and a limit of agreement between 45.0 mg and -58.5 mg. CONCLUSION: Use of the mass quantification algorithm in combination with a calibration phantom allows accurate quantification of coronary calcium. Measurements of calcium mass obtained at 1.25-mm section width CT angiography have the best agreement with those obtained at the traditional 3-mm section width imaging protocol.

Algorithms↗

Coronary artery calcium: accuracy and reproducibility of measurements with multi-detector row CT--assessment of effects of different thresholds and quantification methods.

PURPOSE: To evaluate the effects of different thresholds and quantification methods on the accuracy and reproducibility of coronary calcium measurements with multi-detector row computed tomography (CT). MATERIALS AND METHODS: A cardiac CT phantom containing predetermined calcified cylinders was scanned. Calcium volume and mass were measured at various threshold values ranging from 80 to 230 HU. In 32 patients, two consecutive CT scans were obtained, and the coronary artery calcium score, volume, and mass were measured by one observer at 130- and 90-HU thresholds. Correlation analysis and analysis of variance were performed to evaluate the measurement errors in the phantom study and the interscan variability in the clinical study. RESULTS: In the phantom, mass measurement error varied with threshold and calcium density (P <.01). Mass error was strongly correlated with volume error (r = 0.91, P <.01) but with a much smaller range. In the clinical study, interscan variability of mass measurements was significantly lower than that with other measurement methods for both patients and individual vessels. For the patients, the mean interscan variability of calcium score, volume, and mass at the 130-HU threshold was 20.4%, 13.9%, and 9.3%, respectively. For all methods, interscan variability was not significantly different between the 130- and 90-HU thresholds (P >.05). CONCLUSION: The mass measurement is more accurate, less variable, and more reproducible in coronary calcium quantification than are measurements with other algorithms. Accurate quantification of calcium in each calcified plaque may require that the threshold be set individually, depending on the calcium density.

Algorithms↗

Hemochromatosis: diagnosis and quantification of liver iron with gradient-echo MR imaging.

PURPOSE: To assess the role of magnetic resonance (MR) imaging in detection and quantification of liver iron overload. MATERIALS AND METHODS: MR imaging at 0.5 T was prospectively performed on 77 patients (67 with liver iron overload and 10 without) who underwent a liver biopsy with biochemical determination of the liver iron concentration (LIC) (normal, < 36 mumol per gram of liver tissue [dry weight]). Ratios of signal intensities and liver T2 relaxation time were calculated from images obtained with spin-echo and breath-hold gradient-echo (GRE) sequences. RESULTS: Liver-to-tissue signal intensity ratios were better correlated with LIC than T2 relaxation time. Long-echo-time GRE sequences were the most sensitive for detection of slight overload. Thus, high sensitivity (94%) and specificity (90%) were obtained with a liver-to-fat ratio threshold of 1. The quantification of iron with MR imaging was accurate when the LIC was 80-300 mumol/g. For heavy overload, above 300 mumol/g, quantification was impossible owing to complete signal loss. Pancreatic and splenic signal intensity were unchanged in most cases. CONCLUSION: This method, which can be improved by using more sensitive sequences with a high-field-strength system, should be competitive with biopsy for the diagnosis of substantial liver iron overload.

Female↗

Carotid artery stenosis: clinical efficacy of MR phase-contrast flow quantification as an adjunct to MR angiography.

PURPOSE: To assess the utility of magnetic resonance (MR) flow quantification in the evaluation of suspected carotid artery stenosis. MATERIALS AND METHODS: Fifty-five patients referred for angiography and 10 healthy volunteers underwent Doppler ultrasound, three-dimensional time-of-flight MR angiography, and MR phase-contrast flow quantification to measure peak systolic velocity (PSV) and volumetric flow rate (VFR) in the common and internal carotid arteries distal to the stenosis. RESULTS: PSV and VFR were significantly lower in the severely (> or = 70%) stenosed internal carotid arteries (P < .05). The VFR ratio (internal carotid artery-common carotid artery) achieved 91% overall accuracy for detection of severe stenosis. Combination of flow data with results of MR angiography yielded sensitivity of 100% (95% confidence interval, 78%-100%) with a modest loss in specificity. Doppler and MR measurements of PSV in the common carotid artery showed significant correlation: volunteers, r = .73; patients, r = .64. CONCLUSION: MR flow quantification provides information about the hemodynamic significance of carotid stenosis. As an adjunct to MR angiography, it may be useful in enabling differentiation of occlusions from critical stenoses.

Adult↗

Automatic quantification of immunohistochemically stained cell nuclei based on standard reference cells.

A fully automatic method for quantification of images of immunohistochemically stained cell nuclei by computing area proportions, is presented. Agarose embedded cultured fibroblasts were fixed, paraffin embedded and sectioned at 4 microm. They were then stained together with 4 microm sections of the test specimen obtained from bladder cancer material. A colour based classifier is automatically computed from the control cells. The method was tested on formalin fixed paraffin embedded tissue section material, stained with monoclonal antibodies against the Ki67 antigen and cyclin A protein. Ki67 staining results in a detailed nuclear texture with pronounced nucleoli and cyclin A staining is obtained in a more homogeneously distributed pattern. However, different staining patterns did not seem to influence labelling index quantification, and the sensitivity to variations in light conditions and choice of areas within the control population was low. Thus, the technique represents a robust and reproducible quantification method. In tests measuring proportions of stained area an average standard deviation of about 1.5% for the same field was achieved when classified with classifiers created from different control samples.

Automation↗

Detection and quantification of calcium fluoride using micro-Raman spectroscopy.

Investigations on the caries-preventive effect of calcium fluoride have been seriously hampered by the lack of adequate detection techniques. In this paper, the micro-Raman technique is introduced as a suitable method for CaF2 quantification with a spot size typically 5 microns. Advantages of this measuring technique and associated problems are described for CaF2 determination in the presence of large amounts of hydroxyapatite. The data show that a comparative analysis of Raman intensities of calcium fluoride and phosphate ions of hydroxyapatite can be used for quantification of CaF2 concentrations. The intensity behaviour of the CaF2 and hydroxyapatite Raman lines is explained considering the absorption-scattering cross-sections of the two components. The ratio of the Raman intensity (322 cm-1) of CaF2 to the phosphate Raman intensity (432/447 cm-1) of hydroxyapatite was found to be related linearly to the ratio of the concentration of CaF2 to that of hydroxyapatite. At present, the lower limit of CaF2 quantification is approximately 3 wt%. The technique has already been applied on fluoridated bovine enamel, and Raman emissions due to CaF2 or CaF2-like material formed on/in enamel have been detected for the first time.

Animals↗

A comparison of glottal voice source quantification parameters in breathy, normal and pressed phonation of female and male speakers.

This study concerns a comparison of different quantification techniques that have been developed to parameterize a voice source that has been estimated using inverse filtering that applies no flow mask (i.e. absolute flow values are not obtained). The speech material consisted of voices of three different phonation types produced by 5 female and 5 male subjects. Quantification of the voice source was computed using three time-based quotients that were extracted from the glottal flow waveforms, two time-based parameters that were extracted from the flow and its first derivative, one amplitude-domain quotient that was defined using both the flow and its derivative, and one frequency domain parameter that was computed from the flow signal. The results showed that phonation types could be separated from each other most effectively when quantification was based on parameters that were extracted between the instant of the maximal glottal opening and the minimum peak of the flow derivative.

Female↗

Quantification in situ of crystalline cholesterol and calcium phosphate hydroxyapatite in human atherosclerotic plaques by solid-state magic angle spinning NMR.

Because of renewed interest in the progression, stabilization, and regression of atherosclerotic plaques, it has become important to develop methods for characterizing structural features of plaques in situ and noninvasively. We present a nondestructive method for ex vivo quantification of 2 solid-phase components of plaques: crystalline cholesterol and calcium phosphate salts. Magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectra of human carotid endarterectomy plaques revealed (13)C resonances of crystalline cholesterol monohydrate and a (31)P resonance of calcium phosphate hydroxyapatite (CPH). The spectra were obtained under conditions in which there was little or no interference from other chemical components and were suitable for quantification in situ of the crystalline cholesterol and CPH. Carotid atherosclerotic plaques showed a wide variation in their crystalline cholesterol content. The calculated molar ratio of liquid-crystalline cholesterol to phospholipid ranged from 1.1 to 1.7, demonstrating different capabilities of the phospholipids to reduce crystallization of cholesterol. The spectral properties of the phosphate groups in CPH in carotid plaques were identical to those of CPH in bone. (31)P MAS NMR is a simple, rapid method for quantification of calcium phosphate salts in tissue without extraction and time-consuming chemical analysis. Crystalline phases in intact atherosclerotic plaques (ex vivo) can be quantified accurately by solid-state (13)C and (31)P MAS NMR spectroscopy.

Animals↗

Quantification of perfusion using bolus tracking magnetic resonance imaging in stroke: assumptions, limitations, and potential implications for clinical use.

BACKGROUND: MR techniques have been very powerful in providing indicators of tissue perfusion, particularly in studies of cerebral ischemia. There is considerable interest in performing absolute perfusion measurements, with the aim of improving the characterization of tissue "at risk" of stroke. However, some important caveats relating to absolute measurements need to be taken into account. The purpose of this article is to discuss some of the issues involved and the potential implications for absolute cerebral blood flow measurements in clinical use. SUMMARY OF COMMENT: In bolus tracking MRI, deconvolution of the concentration-time course can in theory provide accurate quantification. However, there are several important assumptions in the tracer kinetic model used, some of which may be invalid in cerebral ischemia. These can introduce significant errors in perfusion quantification. CONCLUSIONS: Although we believe that bolus tracking MRI is a powerful technique for the evaluation of perfusion in cerebral ischemia, interpretation of perfusion maps requires caution; this is particularly true when absolute quantification is attempted. Work is currently under way in a number of centers to address these problems, and with appropriate modeling they may be overcome in the future. In the interim, we believe that it is necessary for users of bolus tracking perfusion data to be aware of the current technical limitations if they are to avoid misinterpretation or overinterpretation of their findings.

Cerebrovascular Circulation↗

Hemodynamic effects of carotid endarterectomy by magnetic resonance flow quantification.

BACKGROUND AND PURPOSE: Blood flow can be evaluated non-invasively using magnetic resonance phase-contrast flow quantification. The purpose of this prospective study was to assess the feasibility of this method and to evaluate the hemodynamic effects of carotid endarterectomy. METHODS: Volumetric flow rates and peak systolic velocities of the internal and common carotid and the vertebral arteries were measured by magnetic resonance flow quantification. Sixteen patients undergoing 18 endarterectomies had complete flow data recorded preoperatively and 3 days after surgery. RESULTS: The inverse correlation between the angiographic stenosis degree and the preoperative flow rate in the corresponding internal carotid artery was highly significant (r = -.69, P < .001). After endarterectomy, the mean flow in the ipsilateral internal carotid artery improved from 143 to 233 mL/min (P < .001). The mean peak systolic velocity increased from 23 to 37 cm/s (P < .001). No significant changes were seen in the contralateral carotid or the vertebral arteries. The mean total blood flow improved by 81 mL/min (P = .08). In the severely stenosed bifurcations (70% to 99%, n = 11), the flow rate improved by 106 mL/min and in the moderately (30% to 69%, n = 4) or mildly (< 30%, n = 3) stenosed bifurcations by 63 mL/min. If the contralateral carotid artery was occluded or severely stenosed, the improvement was 164 mL/min. CONCLUSIONS: Magnetic resonance flow quantification provides a useful tool for the follow-up of the hemodynamic effects of carotid endarterectomy. Our results indicate that surgery is followed by a significant increase of blood flow in the ipsilateral carotid artery and that there appear to be differences in flow increase between subgroups of patients with different degrees of stenosis.

Aged↗

Quantification of bacteria in oral samples by competitive polymerase chain reaction.

Information about the total amount of bacteria in oral samples contributes to assessment of an individual's risk of contracting dental caries or developing periodontitis and the prediction of that individual's clinical course. Since existing techniques are often time-consuming and expensive, it seemed attractive to look for alternative methods for the quantification of eubacteria. With their high specificity and sensitivity, polymerase chain-reaction (PCR) techniques have the potential of supplying fast and reliable results. We developed a method of competitive PCR for the quantification of eubacteria. We designed forward and reverse PCR primers which bind to highly conserved sequences of the bacterial 16S rRNA gene. A homologous competitor was synthesized with Escherichia coli 16S rDNA as a template, with the reverse primer and a hybrid primer which binds 67 bases downstream to the forward primer and carries the forward primer sequence at its 5' end. Specificity controls with 30 different bacterial species, 5 Archaea, 3 fungi, human astrocytoma cells, and rat hepatoblastoma cells were carried out. Results were positive for all eubacteria and negative for all other cells tested. Calibration curves were obtained by co-amplification of known amounts of E. coli cells in the presence of the homologous competitor. The developed method was successfully applied to assessment of the accumulation of bacteria during an oral hygiene cessation experiment. The competitive PCR method proved to be a reliable and fast method for the quantification of bacterial DNA and cultured eubacteria, as well as of bacteria in biological samples. It may find further applications not only in periodontology and cariology but also in other fields of medical microbiology.

Archaea↗

Fluoxetine bioequivalence study: quantification of fluoxetine and norfluoxetine by liquid chromatography coupled to mass spectrometry.

In this study, the authors assessed the bioequivalence of two fluoxetine tablet formulations in 24 healthy volunteers of both sexes who received a single 20 mg dose of each fluoxetine formulation, and a new sensitive method for the quantification of fluoxetine and norfluoxetine in human plasma was developed. The study was conducted using an open, randomized, two-period crossover design with a 4-week washout interval. Plasma samples were obtained over a 672-hour period. Plasma fluoxetine and norfluoxetine concentrations were analyzed by combined liquid chromatography coupled to mass spectrometry (LC-MS) with positive ion electrospray ionization using selected ion recording (SIR). Kolmogorov-Smirnov's test, histograms, probit plots, and the correlation between norfluoxetine AUC(0-infinity) and fluoxetine AUC(0-infinity) were used to analyze the population distribution. The limit of quantification was 0.15 ng.ml-1 and 0.50 ng.ml-1 for both fluoxetine and norfluoxetine, respectively. Within- and between-run imprecision was less than 13% and 17%, respectively. The pharmacokinetic parameters obtained for fluoxetine and norfluoxetine after the administration of each formulation included AUC(0-672 h), AUC(0-infinity), Cmax, Cmax/AUC(0-672 h), tmax, t1/2, and Ke. The AUC values for fluoxetine were not consistent with a normal distribution, reflecting the existence of two different populations (poor and extensive metabolizers). The mean pharmacokinetic parameters for extensive fluoxetine metabolizers were 27.0 ng ml-1 for Cmax, 2064.0 ng h ml-1 for AUC(0-infinity), and 85.4 h t1/2. The mean pharmacokinetic parameters for norfluoxetine (in extensive metabolizers only) were 2532.0 ng h ml-1 for AUC(0-infinity) and 8.4 ng ml-1 for Cmax. For fluoxetine bioequivalence, the 90% CI of the individual ratio geometric mean for Psiquial/Prozac (including both extensive and poor metabolizers) was 101.6% to 121.1% for AUC(0-672 h) and 86.1% to 102.6% for Cmax. For norfluoxetine, the 90% CI of the individual ratio geometric mean for Psiquial/Prozac (including both extensive and poor metabolizers) was 90.3% to 108.3% for AUC(0-672 h) and 84.5% to 106.3% for Cmax. The new method developed (LC-MS) presented high sensitivity, specificity, and short chromatographic run for the quantification of both fluoxetine and norfluoxetine in human plasma. Since both 90% CI for AUC and Cmax geometric mean ratios were included in the 80% to 125% interval proposed by the U.S. Food and Drug Administration, Psiquial was considered bioequivalent to Prozac according to both the rate and extent of absorption. The finding that there were no significant differences in the bioequivalence assessed by either fluoxetine or norfluoxetine pharmacokinetic parameters indicates that future bioequivalence trials may be performed by quantifying fluoxetine only.

Adolescent↗

Analysis of immunohistochemical label of Fos protein in the suprachiasmatic nucleus: comparison of different methods of quantification.

The induction of the proto-oncogene c-fos, and its phosphoprotein product Fos, has been extensively used to study the effects of light on the circadian pacemaker in the suprachiasmatic nucleus (SCN). Experimental approaches to the quantification of Fos induction have mainly been based on immunohistochemistry and subsequent measure of Fos immunoreactivity (IR) in sections of the SCN. In this study, the authors compare several methods of quantification using optical density image analysis or counts of Fos-IR labeled cells. To assess whether optical density measures using image analysis reflect the amount of Fos in brain tissue, the authors developed standards of known concentrations of Fos protein in an agar matrix. The agar standards were sectioned and treated simultaneously with sections of the SCN from animals exposed to different levels of irradiance. Optical density was found to be proportional to the quantity of Fos in the sections, indicating that this measure accurately reflects relative levels of Fos protein induction. Quantification by optical density analysis allows an objective measure in which the various parameters, conditions of illumination, and threshold can be maintained constant throughout the analysis. Counting cells by visual observation is more subjective because threshold values cannot be precisely defined and can vary according to the observer, illumination, degree of label, and other factors. In addition, cell counts involving direct visual observation, automated cell counts, or stereological methods do not take into account the difference in the density of label between cells, thus giving equal weight to lightly or densely stained cells. These measures are more or less weakly correlated with measures of optical density and thus do not accurately reflect the amount of bound Fos protein in the tissue sections. In contrast, labeled surface area as measured by image analysis shows a linear relationship with optical density. The main outcome of this study is that computer-assisted image analysis provides an accurate and rapid method to determine the relative amount of Fos protein in the SCN and the effects of light on intracellular signaling mechanisms involved in the circadian clock.

3,3'-Diaminobenzidine↗

Quantification of chemotactic response of quiescent and proliferating fibroblasts in Boyden chambers by computer-assisted image analysis.

The chemotactic response of human gingival fibroblasts to platelet-derived growth factor (PDGF) was investigated in 48-well modified Boyden chambers. Results were quantified using computer-assisted image analysis of propidium iodidestained cells and were compared with results obtained using the conventional method of quantification by direct counting. Quantification by image analysis was found to be rapid and accurate, and correlated closely with results obtained by direct counting. Bromodeoxyuridine (BrdU)-labeled proliferating cells were double stained with propidium iodide and fluorescein isothiocyanate (FITC)-conjugated anti-BrdU antiserum. The cycling cells showed a markedly reduced chemotactic response to PDGF, whereas cells in S-phase of the cell cycle did not show any response. This method of quantification of results of Boyden chamber assays is simple and reliable, and allows the use of double labels to investigate the chemotactic response of subpopulations of cells within a heterogeneous population of cells.

Cell Cycle↗

The simultaneous quantification of cytochrome P450 dependent linoleate and arachidonate metabolites in urine by HPLC-MS/MS.

A method for the simultaneous quantification of urinary linoleic and arachidonic acid derived epoxides and diols, as well as the arachidonate omega hydroxylated product has been developed. The method employs negative mode electrospray ionization and HPLC with tandem mass spectroscopy for quantification. Odd chain length saturated epoxy and dihydroxy fatty acids are used as analytical surrogates resulting in linear calibrations (r (2) > or = 0.9995). Standard addition analyses showed that matrix effects do not prevent these surrogates from yielding reliable quantitative results. Using 4 ml urine aliquots at a final extract volume of 100 micro l and injecting 10 micro l, method detection limits and limits of quantification were < or =0.5 and 1.5 nM, respectively. The sensitivity for dihydroxy lipids was from 3- to 10-fold greater than the corresponding epoxy fatty acid. Shot to shot run times of 31 min were achieved. Rodent and human urine analyses indicated the method sensitivity is sufficient for general research applications. In addition, diurnal fluctuations in linoleate and arachidonate derived metabolites were observed in human subjects.

Adult↗

Accurate quantification of nonesterified and total cholesterol in plasma and urine from crude lipid extracts by stable isotope dilution and field desorption mass spectrometry.

Levels of free and total cholesterol were quantified in control sera and human urine using [2H7]cholesterol or [13C2]cholesterol as internal standard and field desorption mass spectrometry for the quantification. These analyses are performed using a crude chloroform extract without chromatographic separation of the constituents. Due to the extremely soft ionization conditions in field desorption, the constituents of the crude lipid extract predominantly form molecular ion species and only a very low amount of fragment ions of low relative abundance thus minimizing the possibility of ion overlap in the mass spectrum. Mass spectrometric quantification of cholesterol in two control sera resulted in data within the range of assigned concentrations. The standard deviations of the mass spectrometric data were around +/- 2%. Generally, 50 microliters of serum or plasma or 500 microliters of urine are extracted and the mass spectrometric quantification performed using an aliquot of 2-10%. A characteristic pattern of intact cholesteryl esters is also obtained.

Body Fluids↗

Quantification of low abundance natriuretic peptide receptor mRNA in rat tissues.

Natriuretic peptides are important regulators of vascular resistance and volume and electrolyte homeostasis. The quantification of natriuretic peptide receptor (NPR) mRNA is important for the understanding of the regulation of this humoral system, but is difficult due to low expression of the NPR mRNA. We report here on the evaluation of a polymerase chain reaction (PCR)-aided transcript titration assay for quantification of all three NPR subtypes (NPR-A, NPR-B, and NPR-C) mRNA. A multispecific internal standard RNA with parts of NPR-A, NPR-B, NPR-C and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) nucleotide sequences was constructed and reverse transcription of standard and sample RNA (400 ng) was performed in parallel for all three NPRs and GAPDH. The specific PCR yielded differently sized products, which were quantified by high performance liquid chromatography (HPLC). The determination of specific mRNA concentrations was not influenced by cDNA input and did not depend on the PCR cycle number. Linearity between sample RNA input and mRNA concentration was demonstrated. Application of the evaluated method showed that the NPR-A mRNA expression was the most abundant of the three natriuretic peptide receptor mRNAs in rat lungs, glomeruli and left ventricles, followed by the NPR-C mRNA and the NPR-B mRNA expression. Thus, the described method allows the reliable quantification of the specific mRNA expression of all three NPRs with small amounts of RNA. The presented method might foster future research on the regulation of this humoral system in cardiovascular and kidney diseases.

Animals↗