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October 4, 2005
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October 4, 2005
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October 4, 2005
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July 5, 2005
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Common genetic variants of coagulation factor genes associated with differences in concentration and/or function of coagulation factors have been studied in search of variability that could explain the individual susceptibility to thrombosis and atherothrombotic diseases. The more outstanding polymorphisms in genes of factors involved in coagulation and fibrinolysis described in the literature (such as fibrinogen, factor XIII, glycoprotein IIb/IIIa, von Willebrand factor, factors VII, VIII and IX, factor V, ATIII and protein C system factors, prothrombin, PAI-1 and fibrinolytic system) are reviewed in the context of factor's structure and function and also in its proposed relevance for thrombotic and atherothrombotic risk definition.
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July 5, 2005
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The UF-100™ analyser is a fully automated instrument that stains the DNA and the membranes of the formed elements in native urine. The sample then passes as a laminar flow through a laser beam and light scattering, fluorescence and impedance are measured. The main purpose of the present work was to assess the analytical performance and the accuracy of the measurements of the UF-100™ analyser. No carryover was observed, while the linearity was higher then the upper limit (40000 total particles μl −1 ) suggested by the manufacturer. The within-run imprecision was low, ranging from 17.7 % to 2.4 % and was up to threefold better than manual microscopy. Comparison of results obtained by sediment microscopy (performed according to National Committee for Clinical Laboratory Standards (NCCLS) recommendations) and by the UF-100™ analyser showed a linear correlation with r = 0.833 for erythrocytes, r = 0.934 for leukocytes, r = 0.880 for epithelial cells and r = 0.40 for casts. To evaluate the reliability of the UF-100™ analyser in detecting bacteria we compared the results with the microbial culture (n = 608). Using a cut-off value of bacterial count above 1800 μl −1 and at leukocyte count above 45 μl −1 , the analyser detected positive cultures with a sensitivity of 87 % and a specificity of 80 %. In conclusion, the UF-100™ analyser can improve the work flow, increasing the output of urinalysis by reducing the number of specimens submitted for microscopy. Also the method provides reliable information for the identification of urinary tract inflammation and bacterial infection.
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July 5, 2005
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Urinary microscopy is a diagnostic tool which is largely used by nephrologists. In the opinion of the authors the best results can be achieved when all the aspects concerning this test are properly taken into account. Thus, from the methodological point of view, proper patient guidance, proper urine collection and handling, adequate microscopic equipment, and knowledge of the factors which can influence the results are all necessary. All the elements of clinical importance have to be known, namely, erythrocytes (with their morphological subtypes), leukocytes, tubular cells, uroepithelial cells (both superficial and deep), lipids, casts, crystals, and microorganisms. Then, the urinary findings have to be interpreted and, whenever possible, also combined into urinary profiles (e.g., the nephritic sediment, the nephrotic sediment). This, combined with other laboratory tests, the pathologic findings, and the clinical data, allows for the definition and management of urinary tract diseases.
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July 5, 2005
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The relative density of urine is the ratio of its density to that of water and depends on both the number and weight of solute particles in the sample, while osmolality depends only on the number of solute particles. Water metabolism is regulated by the interaction of the renal medullary countercurrent system with the circulating levels of antidiuretic hormone and thirst. The concentration of solids in urine can be measured by weighing, hydrometry, oscillations of a capillary tube, refractometry and reagent strip. These techniques, interrelated but not identical, are commonly used in hospital laboratories and in clinical wards. We compared the results obtained in 1725 urine samples of inpatients and outpatients using an automated refractometer to those obtained using two visually read dip stick tests. The correlation coefficients (Super Aution analyser vs. Aution Sticks 10 EA, Aution Sticks 10 EA vs. N-Multistix, Super Aution analyser vs. N-Multisticks were 0.663, 0.645 and 0.514, respectively) and the great dispersion of mountain plots demonstrates that different techniques are not interchangeable in the measurement of relative density. Since the results obtained after discarding the samples with pH higher than 7 and those containing glucose or protein were very similar to the ones reported above, the role of these interferents appears negligible in inducing the discrepancy.
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July 5, 2005
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The paper presents a review of the characteristics and analytical performance of the most current dry chemistry methods for detection and estimation of the different urinary proteins. Description of the classical “protein error” dipsticks for macroalbuminuria is supplemented by data on more sensitive alternatives based on the same principle, reaching into the zone of microalbuminuria. Immunological test strips are available for detection of low concentrations of albumin. Application of the same principle is attempted for detection of other specific proteins. Highly sensitive enzymatic reactions allow detection of “lysis proteins”: haemoglobin, leukocyte esterase and β-N-acetyl-glucosaminidase. An efficient strategy for screening for all types of pathological proteinuria based on detection of low albumin levels is presented.
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July 5, 2005
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A quantification of proteins of different molecular size has been shown to be useful in characterizing the mechanism and medical causes of proteinuria. By analyzing urine albumin, α 1 -microglobulin, immunoglobulin G and α 2 -macroglobulin together with total protein, prerenal, glomerular, tubular and postrenal causes of proteinuria can be detected and differentiated by their specific urine protein patterns. Using automated turbidimetric procedures, prerenal proteinurias are characterized by an albumin/total protein ratio below 0.4. Tubulo-interstitial diseases which are negative in the protein test strip procedure are detected and clearly differentiated from other causes of proteinuria by their high α 1 -microglobulin/albumin ratios. In postrenal proteinuria, α 2 -macroglobulin proved to be a useful marker, when albumin excretion exceeds 100 mg/l urine. This protein exhibits plasma-like ratios to albumin in postrenal causes, whereas it is much lower in renal proteinurias. The new strategy, which has been evaluated in more than 500 clinically and partly histologically proven cases of renal diseases, more sensitively detects glomerular and tubulo-interstitial diseases when applied in urine screening and allows us to distinguish all clinically important causes from analysis of a morning spot urine sample.
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July 5, 2005
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Microalbuminuria is still the only early abnormality of the diabetic kidney that has an established prognostic value. Microalbuminuria evolves into clinical nephropathy and renal failure in a majority of cases of insulin-dependent diabetic patients, and is defined by the detection of urinary albumin excretion rates of 20–200 μg/min in timed urine collections. The occurrence of microalbuminuria at rates of 5–27 % of non-proteinuric patients and cost-benefit considerations justify the screening for microalbuminuria in diabetic outpatient clinics. Both near-normalisation of glycaemic control and treatment with ACE-inhibitors are indicated in patients with insulin-dependent diabetes to correct the progression of micro- to macroalbuminuria. Other therapeutic perspectives are being considered, but the current notion that the available therapies may not arrest the course of nephropathy at this stage suggests that earlier interventions may be required. Prevention of microalbuminuria and overt nephropathy may require a primary approach to the subset of patients with a genetic predisposition to this complication, and several studies (candidate gene or genomic scan with microsatellite probes) now address the chromosomal loci and the nature of the genes that may involved.
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July 5, 2005
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A multicentre evaluation of the urine test strip analyser Super Aution-4220 was carried out in six laboratories. The analytical performance of the instrument with regard to imprecision, linearity, detection limit, drift, carry-over and method comparison was studied. Using the Aution stick 8 test strip the pH, glucose, protein, ketones, bilirubin, blood, urobilinogen and leukocyte esterase were analysed. Specific gravity measurements were performed by refractive index method. Within-run and between-run imprecision determined at three levels of analyte were good. No carry-over was observed. Obtained results were linear through all the described analytical range. No significant drift was detected. Method comparison with some quantitative methods was performed and showed a good correlation with most of the analytes. The study of interferences showed minor interferences by common therapeutic drugs with the measurement of some analytes. During the assessment period of about 6 months no breakdown occurred in any laboratory. The Super Aution urine analyser appeared to be a highly automated analyser of urinary test strips. The operation was simple and the maintenance required only a few minutes a day.
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July 5, 2005
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Denaturing gradient gel electrophoresis displays the highest detection rate among mutation scanning methods. In classical denaturing gradient gel electrophoresis the denaturant gradient range and migration times vary for every amplicon to be scanned, greatly affecting the routine application of the method. As an alternative, we developed double gradient denaturing gradient gel electrophoresis where a gradient of pore size is superimposed over the denaturing one, allowing maintenance of the zone-sharpening effect even over prolonged time runs, and adoption of identical run time conditions for all fragments analyzed. Here double gradient denaturing gradient gel electrophoresis has been applied to the analysis of a number of point mutations and polymorphisms located in several exons of three different genes, the cystic fibrosis transmembrane conductance regulator, the β-globin and the p53 genes.
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July 19, 2005
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In the Emergency Department it is mandatory to establish the diagnosis and the prognosis of acute pancreatitis as soon as possible. To evaluate whether the association of serum lipase either with serum β 2 -microglobulin or with C-reactive protein allows simultaneously to establish the diagnosis and the prognosis of acute pancreatitis, 96 patients with acute abdomen were studied. Fifty-eight patients had non-pancreatic acute abdomen and the remaining 38 had acute pancreatitis: 23 mild acute pancreatitis, and 15 severe acute pancreatitis. Forty healthy subjects were studied as controls. Lipase, β 2 -microglobulin and C-reactive protein were determined in the serum of all subjects, using commercial kits. One patient with acute pancreatitis was not correctly classified when lipase was used to discriminate between patients with non-pancreatic acute abdomen and those with acute pancreatitis. For the discrimination of patients with severe acute pancreatitis from those with the mild form of the disease in the remaining 37 acute pancreatitis patients, β 2 -microglobulin had a sensitivity of 53.3 %, specificity of 81.8 %, and prognostic accuracy of 70.3% (27 of the 37 patients correctly classified); 87.5 % of the 96 cases were correctly classified. C-reactive protein showed a lower prognostic accuracy than β 2 -microglobulin: sensitivity 86.7 %, specificity 45.5 %, accuracy 62.2 %; 84.4 % of the cases were correctly classified. Using the polychotomous logistic regression analysis we found the same accuracy in discriminating between patients with acute pancreatitis and those with non-pancreatic acute abdomen (99.0 %) but a lower accuracy (54.1 %) between patients with severe acute pancreatitis and those with the mild form of the disease. Our study shows that the association of serum lipase with β 2 -microglobulin or with C-reactive protein is not useful in simultaneously establishing the diagnosis and prognosis of acute pancreatitis.
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July 19, 2005
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The analytical and clinical performance of a commercial automated immunoassay system (Immulite ® ) for estradiol (E 2 ) in serum was evaluated. The functional sensitivity for E 2 was 0.07 nmol/l, and analytical imprecision (<13 %, <9 % and <7 % at 0.22, 0.51 and 1.51 nmol/l, respectively) for concentrations above this detection limit met published analytical goals. The assay recovery was good and the assay was linear over a wide concentration range. No sample carryover was found, and interferences from common substances present in serum were observed only at very high concentrations. Most of samples from men and postmenopausal women showed E 2 concentrations below the detection limit. Longitudinal estradiol profiles from 11 healthy menstruating women showed characteristic menstrual cycle patterns (12 samples per subject obtained during a 30-day period). Longitudinal studies on women during induction of ovulation showed that E 2 concentrations are highly correlated with the total number of follicles. Our results demonstrate the reliability of this system for routine use in the clinical laboratory.
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July 19, 2005
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Heparin is an effective drug for prevention and treatment of thromboembolic conditions. Although several biological assays have been proposed for monitoring unfractionated heparin therapy, the measurement of the activated partial thromboplastin time (APTT) is the most widely employed test, and the overall risk of thromboembolic episodes was markedly reduced by maintaining APTT ratios above 1.5. However, the adjustment of the heparin therapy on the basis of APTT presents several questions which are still unresolved. Major discrepancies were found in APTTs performed using different reagents in both ex vivo and in vitro heparinized samples and occasionally with different lots of the same reagents; poor correlation was observed between APTT values and plasma heparin concentrations. In order to gain further insights into this phenomenon, we analysed the sensitivity to heparin of five commercial reagents for APTT measurement in 19 ex vivo heparinized samples. Differences were observed; correlation coefficients ranged from 0.820 to 0.985 and slopes of linear regressions from 0.26 to 1.14. Moreover, unsatisfactory correlations were obtained when APTT ratios were compared with heparin plasma concentrations in the same patients' samples. In the heparin therapeutic range of 0.35–0.70 U/ml, reagent-dependent differences were observed in the corresponding APTT values. These results point out a critical role of the assay methodology in monitoring heparin therapy by APTT. We suggest that reference materials and methods should be urgently identified, a universally agreed scale for reporting results should be established and reference ranges for the unfractionated heparin therapy should be reconsidered taking on account the reagent employed.
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July 27, 2005
