Abstract:
BACKGROUND:The genotype of hepatitis C virus (HCV) is a predictor of antiviral therapeutic response. We describe an approach for HCV genotype determination by real-time PCR and melting curve analysis. METHODS:After automated nucleic acid extraction, we used reverse transcription-PCR in a block cycler to amplify nucleotides 6-329 of the 5'-untranslated region of HCV. The product was further amplified by single-tube real-time seminested PCR in a LightCycler instrument (Roche). The final product was analyzed by melting curves with the use of fluorescence resonance energy transfer (FRET) probes. The FRET sensor probe was directed at nucleotides 151-170 of type 1 HCV and was designed to distinguish types 1a/b, 2a/c, 2b, 3a, and 4, with melting temperatures (T(m)s) predicted to differ by 1 degrees C. Genotypes were compared in a blinded fashion with those of the INNO-LiPA(TM) test (Bayer Diagnostics) on 111 serum samples. RESULTS:In preliminary experiments, the Mg(2+) concentration was found to be critical in allowing clear separation of melting points, with the best separation at a Mg(2+) concentration of 2 mmol/L. The results for 111 samples clustered at expected T(m)s for genotypes 1a/b (n = 78), 2a/c (n = 2), 2b (n = 11), 3a (n = 14), and 4 (n = 2). Of the 111 samples, results for 110 were concordant with the comparison method at the level of type 1, 2, 3, or 4. Subtyping results were discordant for two samples, both of type 2. For 108 samples concordant with INNO-LiPA at the genotype and subtype levels, the mean T(m)s were 64.1, 59.5, 54.2, 52.6, and 50.1 degrees C for types 1a/b, 2a/c, 4, 2b, and 3a, respectively, with SDs of 0.2, 0.3, 0.3, 0.2, and 0.3 degrees C. All 78 samples identified as type 1 were concordant with results of the comparison method. CONCLUSIONS:Melting analysis with a single pair of FRET probes can rapidly provide information about HCV genotypes and identifies type 1 samples with high specificity.
journal_name
Clin Chemjournal_title
Clinical chemistryauthors
Bullock GC,Bruns DE,Haverstick DMsubject
Has Abstractpub_date
2002-12-01 00:00:00pages
2147-54issue
12eissn
0009-9147issn
1530-8561journal_volume
48pub_type
杂志文章abstract::In photometry, where both the product formed and one or several reactants absorb light at the same wavelength, the absorbance of the "blank" of the sample at the end of the reaction may be less than that measured at the beginning of the reaction, because of consumption of reactant(s). The blank correction factor for t...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1985-03-01 00:00:00
abstract::Problems encountered in the uroporphyrinogen I synthase assay were found to be due to the Triton reagent, slit-width and excitation wavelength settings of the spectrophotofluorometer, and leg-phase in the enzyme reaction. Of Triton X-100 from four suppliers, only three were found suitable. Differences in enzyme activi...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1980-08-01 00:00:00
abstract:BACKGROUND:The sex differential in coronary heart disease (CHD) risk, which is not explained by male/female differences in lipid and lipoprotein concentrations, narrows with age. We examined whether this differential CHD risk might, in part, be attributable to the sizes of lipoprotein particles or concentrations of lip...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:10.1373/clinchem.2004.032763
更新日期:2004-07-01 00:00:00
abstract::We evaluated a method for quantifying high-density lipoprotein cholesterol in plasma, based on electrophoretic migration of the prestained (with Sudan Black III) sample through a discontinuous polyacrylamide++ gel and densitometric integration of the stain associated with each class of lipoprotein. With this method, o...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1985-11-01 00:00:00
abstract::To study the effect of fatty acid chain saturation on the fluorescence polarization assay as a measure of fetal lung maturity, we used purified phospholipids isolated from human amniotic fluid and various commercial phospholipids. We found that the fluorescence polarization value decreased as the concentration of unsa...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1986-03-01 00:00:00
abstract:INTRODUCTION:Pro or precursor forms of prostate-specific antigen (PSA) have emerged as potentially important diagnostic serum markers for prostate cancer detection. Immunoassays were developed to measure specific proPSA forms containing propeptides of 2, 4, and 7 amino acids [(-2)proPSA, (-4)proPSA, and (-7)proPSA, res...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:10.1373/clinchem.2003.026823
更新日期:2004-06-01 00:00:00
abstract::We measured serum protein and calcium concentrations in 2340 individuals between 10 and 96 years of age from 900 families chosen by probability methods to give a representative population. These values were used to calculate an index, based on a regression analysis of serum protein on calcium, which was then treated a...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1976-10-01 00:00:00
abstract::We describe a novel assay for measuring glycohemoglobin directly from anticoagulated whole blood with the Abbott IMx analyzer. The glycohemoglobin is labeled with a soluble polyanionic affinity reagent and the anionic complex is then captured with a cationic solid-phase matrix. Glycohemoglobin is quantified by measuri...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1993-10-01 00:00:00
abstract::The purpose of Medicare's Demonstration of Competitive Bidding for Clinical Laboratory Services is to determine whether competitive bidding can be used to provide quality laboratory services at prices below current Medicare reimbursement rates. Here, we present key features of the preliminary design for the demonstrat...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1998-08-01 00:00:00
abstract::We describe a liquid-chromatographic assay for AZT in human plasma and urine. This assay involves the use of two internal standards, allowing reference of AZT peaks to the appropriate internal standard, the choice depending on the range of concentrations encountered. This method is isocratic, specific, sensitive enoug...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1988-08-01 00:00:00
abstract::We measured the concentration of carboxyhemoglobin (HbCO) in blood samples from 32 neonates by spectrophotometry (IL282 CO-Oximeter) and gas chromatography, finding a strong positive correlation (r = 0.89) between the concentration of fetal hemoglobin (Hb F) and HbCO as measured by spectrophotometry, but not by gas ch...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1988-05-01 00:00:00
abstract::A method is presented for the assay of delta-aminolaevulinate dehydratase (EC 4.2.1.24) to be performed on 10 microL of whole blood. Results compare favorably with those obtained with an accepted method (J. Clin. Chem. Clin. Bio-chem. 12: 389-390, 1974) that involves use of 200 microL of blood. ...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1981-01-01 00:00:00
abstract::A reaction of urea, o-phthalaldehyde and N-(1-naphthyl)ethylenediamine is described for measurement of urea by manual, continuous-flow, and kinetic methods. The continuous-flow system requires 25 mu-l of sample; 40 samples can be analyzed per hour. The kinetic method requires no enzymes, has no lag phase, and has good...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1975-07-01 00:00:00
abstract:BACKGROUND:Although numerous lateral flow immunoassays (LFIAs) have been developed and widely used, inadequate analytical sensitivity and the lack of multiple protein detection applications have limited their clinical utility. We developed a new LFIA device for the simultaneous detection of high-sensitivity cardiac tro...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:10.1373/clinchem.2011.171694
更新日期:2011-12-01 00:00:00
abstract::We describe a rapid, sensitive, and specific "high performance" liquid chromatographic analysis for disopyramide and its mono-N-dealkylated metabolite in serum, urine, and saliva. We used a mu-Bondapak CN column and an acetate buffer mobile phase containing methanol. Retention times for the two compounds and the inter...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1979-03-01 00:00:00
abstract::We measured total and dialyzable calcium concentrations in consecutive sera submitted for routine lithium analysis. Of 98 samples from 61 different individuals, six (6.1%) total calcium results and 32 (33%) dialyzable calcium results were above the respective reference intervals. By comparison, when both total and dia...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1979-10-01 00:00:00
abstract::We compared a chemiluminescent assay and a colorimetric endpoint assay for measuring an alkaline phosphatase (EC 3.1.3.1) label in an enzyme immunoassay of thyrotropin (TSH). The substrate in the chemiluminescent assay is a derivative of adamantyl 1,2-dioxetane phosphate. On dephosphorylation, catalyzed by alkaline ph...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1989-07-01 00:00:00
abstract::We describe here a nonisotopic immunoassay, based on particle-counting technology, for the determination of urinary albumin. The assay takes only 35 min and has been fully automated on the IMPACT (Acade Diagnostic Systems, Brussels, Belgium) machine. The system measures albumin within a linear range between 6.25 and 5...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1988-02-01 00:00:00
abstract::To determine when a change in serial measurements of prothrombin time in patients receiving oral anticoagulant therapy (ACT) is a statistically significant biological change necessitating dose adjustment, one must know the size of the "critical difference" in statistical terms (i.e., probabilities). In a cohort of 32 ...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1995-03-01 00:00:00
abstract::Creatine kinase isoenzyme BB (CK-BB) is found in the serum of patients with various types of cancer. Using both radioimmunoassay and agarose electrophoresis, we found abnormal amounts of this isoenzyme in the serum of 15 of 17 patients with untreated prostatic carcinoma. Among patients with other types of adenocarcino...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1979-08-01 00:00:00
abstract::Erythrocyte and plasma cholinesterase activities were measured biweekly in one group of 22 subjects for a year and daily for three weeks in another group of nine men. The average range [i.e., (range/mean) X 100] of activity of erythrocyte cholinesterase in men during a year was 8% and during three weeks was 5%. For pl...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1975-12-01 00:00:00
abstract::A new method (enzyme-ligand immunoassay, ELIA) is described for the estimation of estrogen (ER) and progesterone (PR) receptors in microsamples of human breast cancer tissue. The technique, based on the nonisotopic measurement of receptor-bound estradiol and progesterone, involves three steps: (a) simultaneous saturat...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1995-01-01 00:00:00
abstract:BACKGROUND:Measurement of plasma renin is important for the clinical assessment of hypertensive patients. The most common methods for measuring plasma renin are the plasma renin activity (PRA) assay and the renin immunoassay. The clinical application of renin inhibitor therapy has thrown into focus the differences in i...
journal_title:Clinical chemistry
pub_type: 杂志文章,评审
doi:10.1373/clinchem.2008.118000
更新日期:2009-05-01 00:00:00
abstract::We evaluated the HemoCue beta-Glucose Photometer system for use in our neonatal intensive care unit by assaying 178 heparinized whole-blood samples obtained by heel stick. The required sample size is 5 microL. Plasma glucose was analysed by a glucose oxidase/oxygen electrode methodology. Across the glucose range of 1....
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1993-11-01 00:00:00
abstract::A rapid (30-min) semiautomated continuous-flow procedure is described for use in assessing the phospholipids of the particulate ("lamellar body") fraction of human amniotic fluid. The method is based on measuring the difference in fluorescence of 1,6,-diphenyl-1,3,5-hexatriene added to amniotic fluid before and after ...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1983-02-01 00:00:00
abstract::We measured morphine and codeine in commercially available poppy seeds and in serum and urine samples from healthy adults who had ingested these poppy seeds. Four brands of black poppy seeds, examined by gas chromatography-mass spectroscopy (GC-MS) with deuterated internal standards, contained from 17 to 294 microgram...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1987-06-01 00:00:00
abstract::We compared cardiac troponin I (cTnI), using Access, Sanofi Pasteur, and cardiac troponin T (cTnT), using Elecsys, Boehringer Mannheim, in the first two routine blood samplings in a routine panel of cardiac markers for the biochemical diagnostic evaluation of patients with symptoms of acute myocardial infarction (AMI)...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:
更新日期:1998-07-01 00:00:00
abstract:BACKGROUND:Current noninvasive assays for urothelial carcinoma (UC) lack clinical sensitivity and specificity. Given the utility of plasma cell-free DNA (cfDNA) biomarkers, the development of urinary cfDNA biomarkers may improve the diagnostic sensitivity. METHODS:We assessed copy number alterations (CNAs) by shallow ...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:10.1373/clinchem.2019.309633
更新日期:2020-01-01 00:00:00
abstract:BACKGROUND:C-reactive protein (CRP) is a widely recognized indicator of inflammation and is known to play an important role in atherogenesis. Recent prospective studies have demonstrated that increased CRP concentrations within the reference interval are a strong predictor of myocardial infarction, stroke, sudden cardi...
journal_title:Clinical chemistry
pub_type: 杂志文章,评审
doi:10.1373/49.8.1258
更新日期:2003-08-01 00:00:00
abstract:BACKGROUND:There are 45 known genetic diseases that impair the lysosomal degradation of macromolecules. The loss of a single lysosomal hydrolase leads to the accumulation of its undegraded substrates in tissues and increases of related glycoconjugates in urine, some of which can be detected by screening of free oligosa...
journal_title:Clinical chemistry
pub_type: 杂志文章
doi:10.1373/clinchem.2012.201053
更新日期:2013-09-01 00:00:00