Clinical Chemistry
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Role of nitric oxide in cardiovascular disease: focus on the endothelium Abstract
Nitric oxide is a soluble gas continuously synthesized by the endothelium. This substance has a wide range of biological properties that maintain vascular homeostasis, including modulation of vascular dilator tone, regulation of local cell growth, and protection of the vessel from injurious consequences of platelets and cells circulating in blood. A growing list of conditions, including those commonly associated as risk factors for atherosclerosis such as hypertension and hypercholesterolemia, are associated with diminished release of nitric oxide into the arterial wall either because of impaired synthesis or excessive oxidative degradation. Diminished nitric oxide bioactivity may cause constriction of coronary arteries during exercise or during mental stress and contribute to provocation of myocardial ischemia in patients with coronary artery disease. Additionally, diminished nitric oxide bioactivity may facilitate vascular inflammation that could lead to oxidation of lipoproteins and foam cell formation, the precursor of the atherosclerotic plaque. Numerous therapies have been investigated to assess the possibility of reversing endothelial dysfunction by enhancing the release of nitric oxide from the endothelium, either through stimulation of nitric oxide synthesis or protection of nitric oxide from oxidative inactivation and conversion to toxic molecules such as peroxynitrite. Accordingly, causal relationships between improved endothelial function and reduction in myocardial ischemia and acute coronary events can now be investigated.
Clinical Chemistry - Tập 44 Số 8 - Trang 1809-1819 - 1998
The MicroRNA Spectrum in 12 Body Fluids
BACKGROUND
MicroRNAs (miRNAs) are small, noncoding RNAs that play an important role in regulating various biological processes through their interaction with cellular messenger RNAs. Extracellular miRNAs in serum, plasma, saliva, and urine have recently been shown to be associated with various pathological conditions including cancer.
METHODS
With the goal of assessing the distribution of miRNAs and demonstrating the potential use of miRNAs as biomarkers, we examined the presence of miRNAs in 12 human body fluids and urine samples from women in different stages of pregnancy or patients with different urothelial cancers. Using quantitative PCR, we conducted a global survey of the miRNA distribution in these fluids.
RESULTS
miRNAs were present in all fluids tested and showed distinct compositions in different fluid types. Several of the highly abundant miRNAs in these fluids were common among multiple fluid types, and some of the miRNAs were enriched in specific fluids. We also observed distinct miRNA patterns in the urine samples obtained from individuals with different physiopathological conditions.
CONCLUSIONS
MicroRNAs are ubiquitous in all the body fluid types tested. Fluid type–specific miRNAs may have functional roles associated with the surrounding tissues. In addition, the changes in miRNA spectra observed in the urine samples from patients with different urothelial conditions demonstrates the potential for using concentrations of specific miRNAs in body fluids as biomarkers for detecting and monitoring various physiopathological conditions.
Clinical Chemistry - Tập 56 Số 11 - Trang 1733-1741 - 2010
Automated and Manual Direct Methods for the Determination of Blood Urea Abstract
Automated and manual direct methods for the determination of urea in blood or serum are described. These methods determine urea by the colored product formed when urea, in relatively weak acid solution, reacts with diacetyl monoxime in the presence of thiosemicarbazide and ferric ion. Results are compared with those obtained by urease conversion of urea to ammonia and measurement of the ammonia by nesslerization.
Clinical Chemistry - Tập 11 Số 6 - Trang 624-627 - 1965
Design, Construction, and Two Applications for an Automated Flow—Cell Polarization Fluorometer with Digital Read Out: Enzyme-Inhibitor (Antitrypsin) Assay and Antigen—Antibody (Insulin—Insulin Antiserum) Assay Abstract
We have constructed and automated a flow cell polarization fluorometer and demonstrated two specific clinical applications of fluorescence polarization assay, i.e., the enzyme-inhibitor assay of antitrypsin in serum and the antigen-antibody assay of insulin and its antibody. A signal is displayed directly and is immediately available for chart recording and (or) digital data processing. Since fluorescence polarization offers a number of choices in assay parameters and the use of different fluorescent probes, its development for rapid simultaneous measurements of multiple components in body fluids should be possible.
Clinical Chemistry - Tập 19 Số 8 - Trang 838-844 - 1973
Cholesterol in Serum and Lipoprotein Fractions Abstract
1. Methods are described for the separation, by paper electrophoresis and by cold ethanol, of α- and β-lipoproteins in 0.1 ml. of serum, with subsequent analysis of cholesterol in the separated portions.
2. It is shown that both methods of separation yield separated fractions containing substantially the same amounts of cholesterol.
3. Detailed data are given on the errors of measurement for total cholesterol and for cholesterol in the separated lipoprotein fractions.
4. Studies are reported on the stability of cholesterol in stored serum and on paper electrophoresis strips. It is shown that simple drying on filter paper causes no change in cholesterol content and yields a product that is stable for many weeks at ordinary room temperature.
5. The sources of variability in human serum cholesterol values are examined and it is shown that spontaneous intraindividual variability is a much greater source of error than the errors of measurement with these methods.
Clinical Chemistry - Tập 2 Số 3 - Trang 145-159 - 1956
A peroxidase-coupled method for the colorimetric determination of serum triglycerides. Abstract
We describe an enzymatic method for rapid, precise measurement of serum triglycerides with use of sample:reagent ratios as large as 1:200. Hydrolysis of triglycerides is catalyzed by lipase to produce glycerol and free fatty acids. The glycerol generated is then phosphorylated by adenosine 5'-triphosphate in the presence of glycerol kinase. Oxidation of the resulting glycerol 3-phosphate to produce hydrogen peroxide is catalyzed by L-alpha-glycerophosphate oxidase. An intense red chromogen is produced by the peroxidase-catalyzed coupling of 4-aminoantipyrene and sodium 2-hydroxy-3,5-dichlorobenzenesulfonate with hydrogen peroxide. This sensitive chromogen system not only permits use of unusually small sample volumes, it also facilitates a linear response to serum triglyceride concentrations up to at least 10 g/L while displaying good Ringbom (measure of accuracy) characteristics.
Clinical Chemistry - Tập 29 Số 3 - Trang 538-542 - 1983
Dextran sulfate-Mg2+ precipitation procedure for quantitation of high-density-lipoprotein cholesterol.
Clinical Chemistry - Tập 28 Số 6 - Trang 1379-1388 - 1982
Cholesterol in high-density lipoprotein: use of Mg2+/dextran sulfate in its enzymic measurement. Abstract
We describe a method for measuring high-density lipoprotein cholesterol. MgCl2 and dextran sulfate are used to precipitate all low-density and very-low-density lipoproteins. The supernate contains only high-density lipoproteins, the cholesterol concentration of which is estimated by an enzymic method, with a discrete analyzer (Abbott Bichromatic Analyzer). Concentration and instrument response are linearly related to 50 mg/liter. The precision of the method is excellent in the range of clinical interest (100 to 1000 mg of cholesterol per liter). The precision and efficiency of the precipitation are shown at various concentrations of high-density lipoprotein cholesterol. The method was compared to that of two laboratories in the Cooperative Lipoprotein Phenotyping Study group by testing a number of split samples, and agreement was good.
Clinical Chemistry - Tập 24 Số 6 - Trang 931-933 - 1978
Denaturing-HPLC-Based Assay for Detection of ABL Mutations in Chronic Myeloid Leukemia Patients Resistant to Imatinib Abstract
Background: Despite the efficacy of the BCR-ABL tyrosine kinase inhibitor Imatinib mesylate for the treatment of chronic myeloid leukemia (CML), resistance has been observed in a proportion of cases, especially those with advanced stages of the disease. Point mutations within the ABL kinase domain are emerging as the most frequent mechanism for reactivation of kinase activity within the leukemic clone.
Methods: We developed a denaturing-HPLC (D-HPLC)-based assay for screening for ABL point mutations. For each sample, two partially overlapping fragments of 393 and 482 bp corresponding to the kinase domain were amplified by nested reverse transcription-PCR and analyzed under selected temperature and acetonitrile gradient conditions. Fifty-one bone marrow and/or peripheral blood specimens from 27 CML patients who showed cytogenetic resistance to Imatinib were screened in parallel by D-HPLC and by direct sequencing.
Results: In 12 of 27 (44%) patients, D-HPLC showed an abnormal elution profile suggesting the presence of a nucleotide change. Direct sequencing confirmed the presence of a point mutation in all cases. Conversely, all samples scored as wild type by D-HPLC showed no evidence of mutations by direct sequencing. In two cases, novel amino acid substitutions at codons already known for being hot-spots of mutation were identified (F311I and E355D).
Conclusions: The proposed D-HPLC-based assay is highly specific and at least as sensitive as sequencing; with respect to the latter, it provides a much faster and less expensive semiautomated system for mutational screening. It may therefore potentially be a valuable tool for regular, large-scale testing of patients undergoing Imatinib treatment.
Clinical Chemistry - Tập 50 Số 7 - Trang 1205-1213 - 2004
Carnitine determination by an enzymatic cycling method with carnitine dehydrogenase Abstract
We describe a highly sensitive and specific method for determining L-carnitine in serum by use of carnitine dehydrogenase (EC 1.1.1.108). The method involves a new enzymatic cycling technique with NADH, thio-NAD+, and carnitine dehydrogenase, and measures the increase of absorbance at 415 nm of thio-NADH produced at 37 degrees C during the reaction: [formula: see text] The calibration curve for L-carnitine in serum was linear between 5 and 250 mumol/L. Analytical recovery was 96.5-106%, and within-run and between-run imprecisions (CV) were 0.66-4.33% and 1.02-2.56%, respectively. This method was free from interference by bilirubin, hemoglobin, various acyl-DL-carnitines, and ascorbate. The procedure is simple, rapid, accurate, and automatable. The amount of free L-carnitine in serum (53.6 +/- 11.7 mumol/L, n = 200) was greater in men than in women (45.1 +/- 14.2 mumol/L, n = 200) (mean +/- SD).
Clinical Chemistry - Tập 40 Số 5 - Trang 817-821 - 1994
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