bluuebunny/crossref_metadata_2025_split huggingface dataset:
Early Biomarkers of Stroke
Authors: Reynolds Mark A , Kirchick Howard J , Dahlen Jeffrey R , Anderberg Joseph M , McPherson Paul H , Nakamura Kevin K , Laskowitz Daniel T , Valkirs Gunars E , Buechler Kenneth F 
Published: February 2004

Abstract
Background: The diagnosis and management of acute ischemic stroke are limited by the lack of rapid diagnostic assays for use in an emergency setting. Computed tomography (CT) scanning is used to diagnose hemorrhagic stroke but is relatively ineffective (&lt;33% sensitive) in detecting ischemic stroke. The ability to correlate blood-borne protein biomarkers with stroke phenotypes would aid in the development of such rapid tests.
Methods: ELISAs for &gt;50 protein biomarkers were developed for use on a high-throughput robotic workstation. These assays were used to screen plasma samples from 214 healthy donors and 223 patients diagnosed with stroke, including 82 patients diagnosed with acute ischemic stroke. Marker assay values were first compared by univariate analysis, and then the top markers were subjected to multivariate analysis to derive a marker panel algorithm for the prediction of stroke.
Results: The top markers from this analysis were S-100b (a marker of astrocytic activation), B-type neurotrophic growth factor, von Willebrand factor, matrix metalloproteinase-9, and monocyte chemotactic protein-1. In a panel algorithm in which three or more marker values above their respective cutoffs were scored as positive, these five markers provided a sensitivity of 92% at 93% specificity for ischemic stroke samples taken within 6 h from symptom onset.
Conclusion: A marker panel approach to the diagnosis of stroke may provide a useful adjunct to CT scanning in the emergency setting.
Heterogeneity Analysis of the Human Pituitary Proteome
Authors: Zhan Xianquan , Desiderio Dominic M 
Published: February 2004

Abstract
Background: A human proteome is relatively dynamic compared with its corresponding genome. Our aim was to study the heterogeneity of a human pituitary proteome as a function of gender, age, and race.
Methods: Pituitary control tissues (n = 8) were used to extract proteins; each control tissue was analyzed (n = 3–5) with two-dimensional gel electrophoresis (2DGE) and PDQuest software. We obtained 30 high-resolution 2DGE gels and conducted a comparative analysis as a function of gender, age, and race. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry and liquid chromatography-electrospray ionization-quadrupole-ion trap tandem mass spectrometry were used to characterize the protein in each differential spot.
Results: We detected ∼1000 protein spots in each 2DGE map, and 51 differential spots (7 differing with gender, 17 with age, 15 with race, and 12 with the coeffect of age and race). Among those 51, we characterized 28 proteins [5 differing with gender, 8 with age, 6 with race, 8 with the coeffect of age and race, and 1 (somatotropin chain 1) with all of these]. Somatotropin was related to gender, age, and race, and prolactin was higher in females than males. The differentially expressed proteins that were related to age were mainly those proteins associated with cell growth, proliferation, differentiation, apoptosis, and death; those proteins showed no difference with gender and race. Age and race affected some proteins associated with hormone regulation (e.g., follistatin, thyroid hormone receptor β-2, adenylate cyclase-inhibiting Gα protein).
Conclusions: A heterogeneity exists in the human pituitary proteome as a function of gender, age, and race. These findings will serve as a basis for our comparative proteomics studies of human pituitary adenomas.
Blood Test Detecting Autoantibodies to N-Methyl-d-aspartate Neuroreceptors for Evaluation of Patients with Transient Ischemic Attack and Stroke
Authors: Dambinova Svetlana A , Khounteev Guerman A , Izykenova Galina A , Zavolokov Igor G , Ilyukhina Anna Y , Skoromets Alexander A 
Published: February 2004

Abstract
Background: Stroke is a multisystemic disorder that includes mechanisms of thrombosis and neurotoxic coupling. Key metabolites of the molecular cascade following biochemical events appear simultaneously in brain tissue, the blood–brain barrier, and brain vessels, activating the immune system and generating autoantibodies (aAbs) to brain-specific antigens. We developed an ELISA blood test to measure aAbs to a subtype of N-methyl-d-aspartate (NMDA) receptors, which are the key markers of neurotoxicity underlying cerebral ischemia. We investigated the diagnostic accuracy of serum aAbs to NR2A/2B, a subtype of NMDA receptors, in assessing transient ischemic attack (TIA) and ischemic stroke (IS) and its ability to distinguish cerebral ischemia from intracerebral hemorrhage (ICH).
Methods: Autoantibodies to NR2A/2B were measured in 360 serum samples: 105 from TIA/stroke patients and 255 from controls, including patients with controlled hypertension/atherosclerosis and gender- and age-matched healthy individuals.
Results: Patients with TIA (n = 56) and acute IS (n = 31) had significantly higher NR2A/2B aAb concentrations than controls (P &lt;0.0001). The test sensitivities for TIA and IS were 95% and 97%, respectively, and predictive values were 86% and 91% at a cutoff point of 2.0 μg/L. The area under the ROC curve was 0.99. Monitoring NR2A/2B aAbs within 72 h differentiated IS and ICH (P &lt;0.001) and was confirmed by magnetic resonance imaging and computed tomography.
Conclusions: NR2A/2B aAbs are independent and sensitive serologic markers capable of detecting TIA with a high posttest probability and, in conjunction with neurologic observation and neuroimaging, ruling out ICH. The test may help assess risk of TIA in routine general practice and may potentially be useful in assisting diagnosis of acute IS in the emergency setting.
Detecting Subsecond Dopamine Release with Fast-Scan Cyclic Voltammetry in Vivo
Authors: Robinson Donita L , Venton B Jill , Heien Michael L A V , Wightman R Mark 
Published: February 2004

AbstractBackground: Dopamine is a potent neuromodulator in the brain, influencing a variety of motivated behaviors and involved in several neurologic diseases. Measurements of extracellular dopamine in the brains of experimental animals have traditionally focused on a tonic timescale (minutes to hours). However, dopamine concentrations are now known to fluctuate on a phasic timescale (subseconds to seconds).Approach: Fast-scan cyclic voltammetry provides analytical chemical measurements of phasic dopamine signals in the rat brain.Content: Procedural aspects of the technique are discussed, with regard to appropriate use and in comparison with other methods. Finally, examples of data collected using fast-scan cyclic voltammetry are summarized, including naturally occurring dopamine transients and signals arising from electrical stimulation of dopamine neurons.Summary: Fast-scan cyclic voltammetry offers real-time measurements of changes in extracellular dopamine concentrations in vivo. With its subsecond time resolution, micrometer-dimension spatial resolution, and chemical selectivity, it is the most suitable technique currently available to measure transient concentration changes of dopamine.
Multiplex Minisequencing Screen for Common Southeast Asian and Indian β-Thalassemia Mutations
Authors: Wang Wen , Kham Shirley K Y , Yeo Gare-Hoon , Quah Thuan-Chong , Chong Samuel S 
Published: February 2004

AbstractBackground: β-Thalassemia is endemic to many regions in Southeast Asia and India, and &lt;20 β-globin gene mutations account for ≥90% of β-thalassemia alleles in these places. We describe a multiplex minisequencing assay to detect these common mutations.Methods: Gap-PCR was used to simultaneously amplify the β-globin gene from genomic DNA and to detect the Δ619bp deletion mutation. Multiplex minisequencing was then performed on the amplified β-globin fragment to detect an additional 15 common Southeast Asian and Indian β-thalassemia mutations. Site-specific primers of different lengths were subjected to multiple rounds of annealing and single-nucleotide extension in the presence of thermostable DNA polymerase and the four dideoxynucleotides, each labeled with a different fluorophore. Minisequencing products were separated and detected by capillary electrophoresis, followed by automated genotyping. The optimized assay was subjected to a double-blind validation analysis of 89 β-thalassemia and wild-type DNA samples of known genotype.Results: Homozygous wild-type or mutant DNA samples produced electropherograms containing only a single colored peak for each mutation site, whereas samples heterozygous for a specific mutation displayed two different-colored peaks for that mutation site. Samples were automatically genotyped based on color and position of primer peaks in the electropherogram. In the double-blind validation analysis, all 89 DNA samples were genotyped correctly (100% assay specificity).Conclusions: The described semiautomated multiplex minisequencing assay can detect the most common Southeast Asian and Indian β-thalassemia mutations, is amenable to high-throughput scale up, and may bring population-based screening of β-thalassemia in endemic regions a step closer to implementation.
HER-2/neu Gene Copy Number Quantified by Real-Time PCR: Comparison of Gene Amplification, Heterozygosity, and Immunohistochemical Status in Breast Cancer Tissue
Authors: Königshoff Melanie , Wilhelm Jochen , Bohle Rainer M , Pingoud Alfred , Hahn Meinhard 
Published: February 2004

Abstract
Background: Amplification of the oncogene HER-2/neu influences breast cancer pathogenesis, and therapy and prognosis may be affected by the degree of amplification. The extent of amplification or protein overexpression typically is analyzed by fluorescence in situ hybridization or immunohistochemistry (IHC), but quantitative PCR techniques have been described that may provide alternatives to these methods.
Methods: We developed a rapid-cycle, real-time PCR assay for quantification of HER-2/neu gene status. We compared results obtained with this assay with short tandem repeat findings by capillary electrophoresis (CE) and with protein overexpression assessments by IHC. Accuracy and linearity were tested on cell lines and with simulation experiments. We analyzed the amplification of HER-2/neu in 51 clinical tissue samples from patients with suspected breast cancer.
Results: The intra- and interrun CVs for HER-2/neu quantification by real-time PCR were 12% and 18%, and the CV for different simulated amplification and deletion experiments was &lt;7%. The results for HER-2/neu gene status in cell lines matched the values reported in literature. We detected HER-2/neu amplification by real-time PCR in 11 samples, all from patients with invasive ductal carcinoma. Allelic imbalances were found by CE analyses in three samples and by protein overexpression in six samples; five of these were also detected by real-time PCR. Comparison of the quantification results with known prognostic indices yielded results similar to those reported in several other published studies.
Conclusions: The assay is suitable for accurate and precise quantification of HER-2/neu copy numbers in tumor tissue samples obtained in routine clinical practice.
Real-Time Reverse Transcription-PCR Expression Profiling of the Complete Human ATP-Binding Cassette Transporter Superfamily in Various Tissues
Authors: Langmann Thomas , Mauerer Richard , Zahn Alexandra , Moehle Christoph , Probst Mario , Stremmel Wolfgang , Schmitz Gerd 
Published: February 2004

AbstractBackground: ATP-binding cassette (ABC) transporters are involved in many physiologic processes, such as lipid transport, sterol homeostasis, immune mechanisms, and drug transport, and cause various human inherited diseases. Thus, the analysis of ABC transporter mRNA expression profiles for basic research, especially in the field of lipid metabolism, for clinical diagnosis, and for monitoring of drug effects is of great interest.Methods: We have developed a rapid, accurate, and highly sensitive real-time reverse transcription-PCR (RT-PCR) method for detection and quantification of all 47 currently known members of the ABC transporter superfamily. Our expression analysis is based on relative quantification using a calibration curve method. With our assay, expression monitoring of a large number of RNA samples in a 384-well format with only 50 ng of total RNA is possible.Results: In contrast to previous expression analyses of single ABC genes, our method allows the rapid and complete analysis of all ABC transporters in given RNA samples. We used our newly established expression panel to study the gene expression of all human ABC transporters in 20 different human tissues. As a result, we identified tissues with high transcriptional activity for ABC transporters. These organs are mainly involved in secretory function (adrenal gland), metabolic function (liver), barrier function (lung, trachea, small intestine), and tropic function (placenta, uterus).Conclusions: Our RT-PCR assay allows rapid, high-throughput transcriptional profiling of the complete ABC transporter superfamily and thus provides a new enabling tool for research, clinical diagnosis of disease, and drug testing and development.
Minor Increases in Plasma Troponin I Predict Decreased Left Ventricular Ejection Fraction after High-Dose Chemotherapy
Authors: Sandri Maria Teresa , Cardinale Daniela , Zorzino Laura , Passerini Rita , Lentati Paola , Martinoni Alessandro , Martinelli Giovanni , Cipolla Carlo Maria 
Published: February 2004

Abstract
Background: Increased cardiac troponin I (cTnI) in patients treated with high-dose chemotherapy (HDCT) for aggressive malignancy has been proposed as an early marker of late HDCT-induced cardiac dysfunction. We investigated whether cTnI measured by the Stratus CS (Dade Behring) would allow detection of minimal cTnI increases in patients treated with HDCT.
Methods: Plasma cTnI concentrations were determined in 179 consecutive patients before HDCT, at the end of the treatment, and after 12, 24, 36, and 72 h. Cardiac function was explored by echocardiography, and left ventricular ejection fraction (LVEF) was recorded during follow-up. The greatest variation in LVEF from the baseline value was used as a measure of cardiac damage.
Results: In 99 healthy volunteers, the 99th percentile was at 0.07 μg/L. On the basis of ROC curve analysis (area under the curve, 0.89), a cutoff of 0.08 μg/L was chosen (sensitivity, 82%; specificity, 77%). cTnI ≥0.08 μg/L occurred in 57 patients (32%) with echocardiographic monitoring revealing a mean decrease in LVEF of 18%. In comparison, the group of cTnI-negative patients had a mean decrease in LVEF of 2.5% (P &lt;0.001).
Conclusions: Plasma cTnI, as measured with the Stratus CS, can detect minor myocardial injury in patients treated with HDCT.
Differences in Nucleotide Hydrolysis Contribute to the Differences between Erythrocyte 6-Thioguanine Nucleotide Concentrations Determined by Two Widely Used Methods
Authors: Shipkova Maria , Armstrong Victor William , Wieland Eberhard , Oellerich Michael 
Published: February 2004

AbstractBackground: Measurement of 6-thioguanine nucleotide (6-TGN) concentrations in erythrocytes is widely accepted for use in optimization of thiopurine therapy. Various chromatographic methods have been developed for this purpose. In preliminary experiments we observed a considerable difference between 6-TGN concentrations determined with two widely used methods published by Lennard (Lennard L. J Chromatogr 1987;423:169–78) and by Dervieux and Boulieu (Dervieux T, Boulieu R. Clin Chem 1998;44:551–5). We therefore investigated methodologic differences between the two procedures with respect to hydrolysis of 6-TGNs to 6-thioguanine (6-TG) in more detail.Methods: We analyzed 6-TGNs in erythrocyte preparations (n = 50) from patients on azathioprine therapy by both methods, using the original protocols. In one set of experiments, we replaced the 0.5 mol/L sulfuric acid in the Lennard method with the 1 mol/L perchloric acid used by Dervieux and Boulieu. In a second set of experiments, we investigated the effect of various dithiothreitol (DTT) concentrations on 6-TG recovery with both methods. In a third set of experiments, we determined the effect of hydrolysis time on both protocols.Results: Direct comparison of both methods showed that 6-TGN concentrations were, on average, 2.6-fold higher in the Dervieux–Boulieu method over the concentration range tested, although the correlation (r = 0.99; P &lt;0.001) was good. Replacement of sulfuric acid by perchloric acid reduced this difference to ∼1.4-fold (r = 0.99; P &lt;0.001). Increasing the DTT concentration enhanced 6-TG recovery. The hydrolysis time used in the Lennard method (1 h) was not sufficient to achieve complete hydrolysis.Conclusions: The difference between 6-TGN concentrations measured by the two methods is attributable, at least in part, to differences in the extent of nucleotide hydrolysis. For optimization of thiopurine therapy, method-dependent therapeutic ranges are necessary, which precludes comparison of results from clinical studies derived with these methods. Efforts must therefore be made to standardize the analytical procedures for the determination of 6-TGN.
Determination of Choline, Betaine, and Dimethylglycine in Plasma by a High-Throughput Method Based on Normal-Phase Chromatography–Tandem Mass Spectrometry
Authors: Holm Pål I , Ueland Per Magne , Kvalheim Gry , Lien Ernst A 
Published: February 2004

Abstract
Background: The quaternary ammonium compounds, choline and betaine, and dimethylglycine (DMG) reside along a metabolic pathway linked to the synthesis of neurotransmitters and membrane phospholipids and to homocysteine remethylation and, therefore, folate status. Lack of a convenient, high-throughput method for the determination of these compounds has prevented population-based studies of their possible associations with lifestyle, nutrition, and chronic diseases.
Methods: Serum or plasma samples were deproteinized by mixing with three volumes of acetonitrile that contained d9-choline and d9-betaine as internal standards. We used a normal-phase silica column for the separation of choline (retention time, 2.8 min), betaine (1.3 min), DMG (1.15 min), and internal standards, which were detected as positive ions by tandem mass spectroscopy in the multiple-reaction monitoring mode, using the molecular transitions m/z 104→60 (choline), m/z 113→69 (d9-choline), m/z 118→59 (betaine), m/z 127→68 (d9-betaine), and m/z 104→58 (DMG).
Results: For all three metabolites, the assay was linear in the range 0.4–400 μmol/L, and the lower limit of the detection (signal-to-noise ratio = 5) was ≤0.3 μmol/L. The within- and between-day imprecision (CVs) was 2.1–7.2% and 3.5–8.8%, respectively. The analytical recovery was 87–105%. The fasting plasma concentrations (median, 25th–75th percentiles) were 8.0 (7.0–9.3) μmol/L for choline, 31.7 (27.0–41.1) μmol/L for betaine, and 1.66 (1.30–2.02) μmol/L for DMG in 60 healthy blood donors. In individuals who had eaten a light breakfast, plasma concentrations of all three metabolites were significantly (25–30%) higher than in fasting individuals.
Conclusion: This is the first method for the combined measurement of choline, betaine, and DMG in human plasma or serum. The assay is characterized by simple sample preparation, no derivatization, high throughput, imprecision (CV) &lt;10%, detection limits below the values seen in volunteers, and the high specificity provided by tandem mass spectroscopy.
Effect of Riboflavin Status on the Homocysteine-lowering Effect of Folate in Relation to the MTHFR (C677T) Genotype
Authors: Moat Stuart J , Ashfield-Watt Pauline A L , Powers Hilary J , Newcombe Robert G , McDowell Ian F W 
Published: February 2004

Abstract
Background: Riboflavin (vitamin B2) is the precursor for FAD, the cofactor for methylenetetrahydrofolate reductase (MTHFR). MTHFR catalyzes the formation of 5-methyltetrahydrofolate, which acts as a methyl donor for homocysteine remethylation. Individuals with the MTHFR 677C→T mutation have increased plasma total homocysteine (tHcy) concentrations, particularly in association with low folate status. It has been proposed that riboflavin may act together with folate to lower plasma tHcy, particularly in individuals with the thermolabile MTHFR T variant.
Methods: We measured B-vitamin status and plasma tHcy in 126 healthy individuals 20–63 years of age (42 CC, 42 CT, and 42 TT MTHFR genotypes) at baseline and after three interventions (4 months): placebo plus natural diet; daily 400 μg folic acid supplement plus natural diet; and increased dietary folate to 400 μg/day.
Results: At baseline and after nutritional intervention, lower riboflavin status was associated with increased plasma tHcy concentrations. Plasma tHcy was 2.6 μmol/L higher in the lowest plasma riboflavin quartile compared with the highest (P &lt;0.02) and was 4.2 μmol/L higher in the highest erythrocyte glutathione reductase activation coefficient (EGRAC) quartile compared with the lowest (P &lt;0.001). This effect was not restricted to those with the T allele. Folic acid given as a 400 μg/day supplement appeared to exacerbate a tendency toward riboflavin deficiency, as suggested by an increase in the proportion of individuals with EGRAC ≥1.4 from 52% to 65% after supplementation (P &lt;0.05).
Conclusions: Folate and riboflavin interact to lower plasma tHcy, possibly by maximizing the catalytic activity of MTHFR. The effect may be unrelated to MTHFR genotype.
National Academy of Clinical Biochemistry Laboratory Medicine Practice Guidelines: Recommendations for the Use of Laboratory Tests to Support Poisoned Patients Who Present to the Emergency Department
Authors: Wu Alan H B , McKay Charles , Broussard Larry A , Hoffman Robert S , Kwong Tai C , Moyer Thomas P , Otten Edward M , Welch Shirley L , Wax Paul 
Published: February 2004

AbstractBackground: Exposure to drugs and toxins is a major cause for patients’ visits to the emergency department (ED).Methods: Recommendations for the use of clinical laboratory tests were prepared by an expert panel of analytical toxicologists and ED physicians specializing in clinical toxicology. These recommendations were posted on the world wide web and presented in open forum at several clinical chemistry and clinical toxicology meetings.Results: A menu of important stat serum and urine toxicology tests was prepared for clinical laboratories who provide clinical toxicology services. For drugs-of-abuse intoxication, most ED physicians do not rely on results of urine drug testing for emergent management decisions. This is in part because immunoassays, although rapid, have limitations in sensitivity and specificity and chromatographic assays, which are more definitive, are more labor-intensive. Ethyl alcohol is widely tested in the ED, and breath testing is a convenient procedure. Determinations made within the ED, however, require oversight by the clinical laboratory. Testing for toxic alcohols is needed, but rapid commercial assays are not available. The laboratory must provide stat assays for acetaminophen, salicylates, co-oximetry, cholinesterase, iron, and some therapeutic drugs, such as lithium and digoxin. Exposure to other heavy metals requires laboratory support for specimen collection but not for emergent testing.Conclusions:Improvements are needed for immunoassays, particularly for amphetamines, benzodiazepines, opioids, and tricyclic antidepressants. Assays for new drugs of abuse must also be developed to meet changing abuse patterns. As no clinical laboratory can provide services to meet all needs, the National Academy of Clinical Biochemistry Committee recommends establishment of regional centers for specialized toxicology testing.