Abstract：
BACKGROUND:Multiplexed amplicon melting is a closed-tube method for genotyping that does not require probes, real-time analysis, asymmetric PCR, or allele-specific PCR; however, correct differentiation of homozygous mutant and wild-type samples by melting temperature (T(m)) analysis requires high-resolution melting analysis and controlled reaction conditions. METHODS:We designed 4 amplicons bracketing the F5 [coagulation factor V (proaccelerin, labile factor)] 1691G>A, MTHFR (NADPH) 1298A>C, MTHFR 677C>T, and F2 [coagulation factor II (thrombin)] 20210G>A gene variants to melt at different temperatures by varying amplicon length and adding GC- or AT-rich 5' tails to selected primers. We used rapid-cycle PCRs with cycles of 19-23 s in the presence of a saturating DNA dye and temperature-correction controls and then conducted a high-resolution melting analysis. Heterozygotes were identified at each locus by curve shape, and homozygous genotypes were assigned by T(m). We blinded samples previously genotyped by other methods before analysis with the multiplex melting assay (n = 110). RESULTS:All samples were correctly genotyped with the exception of 7 MTHFR 1298 samples with atypical melting profiles that could not be assigned. Sequencing revealed that these 5 heterozygotes and 2 homozygotes contained the unexpected sequence variant MTHFR 1317T>C. The use of temperature-correction controls decreased the T(m) SD within homozygotes by a mean of 38%. CONCLUSION:Rapid-cycle PCR with high-resolution melting analysis allows simple and accurate multiplex genotyping to at least a factor of 4.
journal_name
Clin Chemjournal_title
Clinical chemistryauthors
Seipp MT,Pattison D,Durtschi JD,Jama M,Voelkerding KV,Wittwer CTdoi
10.1373/clinchem.2007.097121subject
Has Abstractpub_date
2008-01-01 00:00:00pages
108-15issue
1eissn
0009-9147issn
1530-8561pii
clinchem.2007.097121journal_volume
54pub_type
杂志文章abstract：:Ethchlorvynol is a commonly abused sedative hypnotic. I report here a seven-day study of serum ethchlorvynol concentrations in a patient who had received an extreme overdose. These concentrations were correlated with the patient's response to stimuli. Serum ethchlorvynol concentrations were logarithmically related to ...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1975-06-01 00:00:00
abstract：:A new instrument has been developed that is similar in design and concept to the GeMSAEC centrifugal analyzer (Oak Ridge National Laboratory) except that electrochemical detection is used rather than optical detection. The present version has eight channels, each with its own detector. Problems of speed control and sa...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1982-09-01 00:00:00
abstract：BACKGROUND:We investigated agreement between self-reported prenatal alcohol exposure (PAE) and objective meconium alcohol markers to determine the optimal meconium marker and threshold for identifying PAE. METHODS:Meconium fatty acid ethyl esters (FAEE), ethyl glucuronide (EtG), and ethyl sulfate (EtS) were quantified...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2014.233718
更新日期：2015-03-01 00:00:00
abstract：BACKGROUND:An assay using a mouse antisialyl Lewis X (sLeX) antibody (CSLEX-1) is used clinically for screening and monitoring patients with breast cancer in Japan. However, the IgM isoform of CSLEX-1 is not preferred for the assay because the bulkiness of IgM generally causes poor accessibility to the antigen. To solv...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2016.260968
更新日期：2016-11-01 00:00:00
abstract：BACKGROUND:Cardiovascular mortality is markedly higher in Russia than in Western Europe and the US. Little is known about indicators of atherosclerotic risk in the Russian population. To our knowledge, this is the first study of apolipoprotein (apo) AI and B in Russia based on the WHO-IFCC standard. METHODS:We measure...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2003.023853
更新日期：2004-02-01 00:00:00
abstract：BACKGROUND:New genetic forms of hereditary hemochromatosis (HH) or hereditary hyperferritinemia (HF) have been identified over the last few years, and abnormalities of various genes may interact in a single patient. This study aimed to develop a rapid automated method for sequencing the main genes involved. METHODS:We...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2007.090605
更新日期：2007-12-01 00:00:00
abstract：:We have assessed the technical performance and robustness of NycoCard CRP Whole Blood, a near-patient test for C-reactive protein (CRP), when used in realistic daily routine situations in general practice clinics (GPC). Thirteen GPCs participated, five of them with technician staff. From 898 patients, split-sample mea...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1997-11-01 00:00:00
abstract：:Investigating the interference of danazol in testosterone radioimmunoassays, we measured serum testosterone concentrations in the serum of women receiving danazol, with three testosterone radioimmunoassay kits. All of these showed positive interference by danazol or its metabolites (or both). Values for testosterone o...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1983-01-01 00:00:00
abstract：:Using image-analyzing equipment, we measured the effect of quinidine on the activity of naphthol AS-D chloroacetate esterase, alpha-naphthyl butyrate esterase, alpha-naphthyl acetate esterase, and acid phosphatase in individual cells of a human Hep G2 hepatoma cell line. The impact of the drug on the morphology of the...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1988-03-01 00:00:00
abstract：BACKGROUND:Standardization of clinical measurements of C-reactive protein (CRP) is based on the availability of reference materials. The serum protein reference material ERM-DA470 was used as a calibrant for various commercially available immunoassays but has now been exhausted. The recently released ERM-DA470k/IFCC wa...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2010.147124
更新日期：2010-09-01 00:00:00
abstract：BACKGROUND:Variability among assays used to measure intact parathyroid hormone (iPTH) is of particular concern because of the routine use of iPTH assay results to guide management of osteodystrophy and calcium metabolism in patients with end-stage renal disease (ESRD). The aim of this study was to determine the extent ...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2006.071589
更新日期：2006-09-01 00:00:00
abstract：BACKGROUND:Although lipoprotein-associated phospholipase A(2) (Lp-PLA(2)) levels are associated with cardiovascular events, Lp-PLA(2) is physically linked to LDL cholesterol (LDL-C). Whether measures of Lp-PLA(2) mass or activity continue to predict risk after LDL-C reduction by statin therapy is uncertain. METHODS:Lp...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2011.180281
更新日期：2012-05-01 00:00:00
abstract：:We measured immunoglobulins in the sera of 33 patients on days 1, 3, 6, 10, and 17 and three to four weeks after surgical operations (mostly hysterectomy or appendectomy) or (six patients) after spinal injury. In the absence of infection or blood transfusion, IgG usually decreased slightly and transiently after hyster...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1975-05-01 00:00:00
abstract：:Measurement of acetaminophen by analysis of the second derivative of its spectrum is specific and sensitive. The method of extraction and the use of just one phosphate buffer as reagent makes this method very convenient. Readings are reliable from 10 to 1500 mg/L. A turnaround time of 20 min makes this method well sui...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1988-06-01 00:00:00
abstract：:Monoclonal antibody CKM-G01 inhibited greater than 99% of the activity of porcine and human creatine kinase(CK)-MM isoenzyme purified from muscle. However, it inhibited only 54% of CK-MM in human serum. Chromatofocusing of serum CK-MM showed that CKM-G01 inhibited 100% of MM3 but not isoform MM1. CKM-G01 inhibited CK-...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1990-01-01 00:00:00
abstract：:A highly sensitive two-site enzyme immunoassay (Capcellia) was developed to determine the concentration of CD4 and CD8 molecules expressed on the surface of human T lymphocytes. This assay, performed in one step (20 min), involves the specific immunocapture of T lymphocytes and reaction of the CD4 or CD8 molecules wit...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1994-01-01 00:00:00
abstract：BACKGROUND:Vascular age is an emerging health indicator and predictor of end-organ damage to the heart, brain, and kidney. Although there have been many review publications concerning risk factors for vascular aging, most include cross-sectional epidemiological studies, limiting inferences about temporality. There is a...
journal_title：Clinical chemistry
pub_type： 杂志文章,评审
doi：10.1373/clinchem.2018.287623
更新日期：2019-01-01 00:00:00
abstract：:We describe a discrete automated radioimmunoassay system for determining choriomammotropin (placental lactogen) in human serum. With the present system it can be measured in as many as 37 unknown sera (50 muL) and three quality-control sera, in duplicate, within 1.5 h. The time required for sample preparation, incubat...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1979-01-01 00:00:00
abstract：:We describe a new method for quantitatively measuring substances of clinical interest by high-performance liquid affinity chromatography (HPLAC). As a model system we selected analysis for transferrin in human serum with immobilized antibodies in a high-performance liquid chromatographic system. SelectiSpher-10 Activa...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1988-10-01 00:00:00
abstract：BACKGROUND:The "unit" for "enzymic activity" (U = 1 micromol/min) was recommended by the International Union of Biochemistry and Molecular Biology (IUB) in 1961 and is widely used in medical laboratory reports. The general trend in metrology, however, is toward global standardization through defining units coherent wit...
journal_title：Clinical chemistry
pub_type： 历史文章,杂志文章
doi：
更新日期：2002-03-01 00:00:00
abstract：BACKGROUND:Although it is being increasingly applied, blood collection for drug testing in sport presents some logistic issues that complicate full applicability on a large scale. The use of dried blood spots (DBS) could benefit compliant blood testing considerably owing to its simplicity, minimal invasiveness, analyte...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2016.257592
更新日期：2016-10-01 00:00:00
abstract：BACKGROUND:Blood concentrations of the calcineurin inhibitors (CNIs) cyclosporine and tacrolimus are currently measured to monitor immunosuppression in transplant patients. The measurement of calcineurin (CN) phosphatase activity has been proposed as a complementary pharmacodynamic approach. However, determining CN act...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/clinchem.2013.213264
更新日期：2014-02-01 00:00:00
abstract：:This rapid radioimmunoassay for aldosterone is performed directly on 100 microL of unprocessed plasma, with 125I-labeled aldosterone as the labeled antigen. Our use of steroid-free plasma in preparing the standard curve resulted in an overestimate of aldosterone; this problem was overcome by adding to such plasma a mi...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1983-02-01 00:00:00
abstract：:We developed a solid-phase two-site immunoenzymometric assay (IEMA) of the estrogen-induced 52-kDa cathepsin D (EC 3.4.23.5) and its processed forms (48-kDa and 34-kDa proteins) in cytosols of breast cancer tissues, using two monoclonal antibodies directed against two different epitopes of these antigens. The first an...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1989-01-01 00:00:00
abstract：:We review the following labels used as substitutes for radioisotopes in immunoassay systems: bacteriophages, chemiluminescence precursors, fluorochromes, fluorogens, fluorescence quenchers, enzymes, coenzymes, inhibitors, substrates, various particulates, metal atoms, and stable free radicals. New methods for performi...
journal_title：Clinical chemistry
pub_type： 杂志文章,评审
doi：
更新日期：1981-07-01 00:00:00
abstract：:We evaluated the Reflotron (Boehringer Mannheim) system for measuring activities of alanine aminotransferase (EC 2.6.1.2) and gamma-glutamyltransferase (EC 2.3.2.2). Within- and between-day precision were comparable with conventional analytical methods. The linear range of the methods extended to more than 40-fold the...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1987-06-01 00:00:00
abstract：BACKGROUND:The detection of monoclonal free light chains (FLCs) is an important diagnostic aid for a variety of monoclonal gammopathies and is especially important in light-chain diseases, such as light-chain myeloma, primary systemic amyloidosis, and light-chain-deposition disease. These diseases are more prevalent in...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：2002-09-01 00:00:00
abstract：:We describe a modification of the MacGee method [Anal. Chem. 42, 421 (1970)] for rapid determination of phenytoin (diphenylhydantoin) in plasma by gas-liquid chromatography, in which an ethylaing reagent is used instead of the more common methylating reagents. With this modification, several N-methylated antiepileptic...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1975-05-01 00:00:00
abstract：:Radioimmunoassay of triiodothyronine was performed with a homogeneous incubation system. Bound and free ligand were separated by partitioning in an aqueous two-phase system. The agreement between the results obtained with this technique and conventional heterogeneous radioimmunoassay was very good (r = 0.981). ...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：
更新日期：1982-04-01 00:00:00
abstract：BACKGROUND:Laboratory tests can be an important source of blood loss in hospitals, especially for newborns and patients in intensive care. The aim of this study was to quantify blood loss for laboratory diagnostic tests in a large number of patients in a teaching hospital. METHODS:We estimated blood loss by multiplyin...
journal_title：Clinical chemistry
pub_type： 杂志文章
doi：10.1373/49.10.1651
更新日期：2003-10-01 00:00:00
