bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular diagnosis of Charcot-Marie-Tooth 1A disease and hereditary neuropathy with liability to pressure palsies by quantifying CMT1A-REP sequences: consequences of recombinations at variant sites on chromosomes 17p11.2-12
Authors: Vandenberghe A , Latour P , Chauplannaz G , Chapon F , Pouget J , Dumas R , Laguenay A , Ollagnon E , Bost M , Duthel S , Chazot G , Boucherat M 
Published: January 2020

Abstract
The most frequent form of Charcot-Marie-Tooth disease (CMT1A; OMIM118.220) is the result of a duplication on chromosome 17 in pll.2-p12. This region contains PMP22, a gene expressed in peripheral myelin. The mutation results from an unequal crossing-over involving repeated sequences, CMT1A-REP, located on both sides of the duplicated region. The reciprocal product of this recombination is a deletion of the same region, which is associated with hereditary neuropathy with liability to pressure palsies (HNPP; OMIM162.500). Proximal and distal CMT1A-REP sequences can be distinguished by the presence of a variant EcoRI site. We quantified the number of these repeat sequences in 36 CMT1A and 40 HNPP patients. CMT1A-REP sequences are involved in almost all of the mutations. The majority of recombination breakpoints occur distally from the variant EcoRI site. However, a few have a breakpoint proximal to this site, which creates the risk of misinterpretation with respect to a duplicated/deleted status.
Analytical performance and clinical validity of two free prostate-specific antigen assays compared
Authors: Jung K , Stephan C , Lein M , Henke W , Schnorr D , Brux B , Schürenkämper P , Loening S A 
Published: January 2020

Abstract
We compared two recently introduced commercial assays (CanAg and Immulite) for measuring free prostate-specific antigen (f-PSA), total PSA (t-PSA), and the ratio of t-PSA/f-PSA (f-PSA%) in control materials and sera of 54 healthy men, 50 patients with benign prostatic hyperplasia (BPH), and 45 patients with prostate cancer (PCa). The lower detection limits for f-PSA were 0.038 microgram/L and 0.004 microgram/L for the CanAg and Immulite assays, respectively. The within-run and between-day precisions of the Immulite assay were &lt; 5%; the CanAg assay showed a poorer precision. Whereas f-PSA values differed between controls and patients but not between BPH and PCa patients, the f-PSA% values were lower in PCa patients than in BPH patients and controls. The receiver-operating characteristic (ROC) curve showed an improved diagnostic power of f-PSA% compared with t-PSA to discriminate between BPH and PCa. Discrimination limits of 16% (CanAg assay), and 15% (Immulite assay) are recommended for f-PSA%.
Long-term stability of blood glutathione and cysteine in humans
Authors: Richie J P , Abraham P , Leutzinger Y 
Published: January 2020

Abstract
Glutathione (GSH) availability is an important factor in the maintenance of health. Accordingly, blood GSH has been proposed as an indicator of health status. To validate the use of blood GSH in population studies, we investigated the long-term intraindividual variation of blood GSH and cyst(e)ine (Cys and cystine) concentrations in healthy adults. In a longitudinal study of 10 subjects, GSH and cyst(e)ine were measured in blood samples collected weekly over 8 months. The average within-person CV for GSH was only 9.1% compared with an observed interindividual CV of 20%. Blood cyst(e)ine was more variable within individuals (mean CV 14.7%) than GSH, whereas the interindividual CV for cyst(e)ine was lower (8.6%). The results demonstrate the stability of blood GSH in free-living subjects and the utility of GSH measurements as an indicator of long-term GSH status.
High sensitivity of the single-strand conformation polymorphism method for detecting sequence variations in the low-density lipoprotein receptor gene validated by DNA sequencing
Authors: Jensen H K , Jensen L G , Hansen P S , Faergeman O , Gregersen N 
Published: January 2020

Abstract
We designed oligonucleotide primer pairs to amplify the promoter region, the translated exon sequences, and the flanking intron sequences of all 18 exons of the LDL receptor gene to compare the ability of the PCR single-strand conformation polymorphism (PCR-SSCP) method with semiautomated solid-phase genomic DNA sequencing to detect sequence variations. In 20 apparently unrelated Danish patients with a clinical diagnosis of heterozygous familial hypercholesterolemia (FH), we identified 13 different mutations in the LDL receptor gene: two silent (C331C, N494 N); five missense (W66G, E119K, T383P, W556S, T7051); one nonsense (W23X); three splice-site (313 + 1G--&gt;A, 1061-8T--&gt;C, 1846-1G--&gt;A); and two frameshift (335del10, 1650delG) mutations. Four of these mutations, N494 N, T383P, 1061-8T--&gt;C, and W556S, have not been reported earlier. The pathogenicity of the T383P, 1061-8T--&gt;C, and W556S mutations remains to be established by in vitro mutagenesis and transfection studies. One patient had three mutations (335del10, 1061-8T--&gt;C, and T705I) on the same allele. Further, nine well-known polymorphisms were detectable with this methodological setup. Direct DNA sequencing of the PCR products used for the SSCP analysis did not reveal any sequence variations not detected by the PCR-SSCP method. In two patients we did not detect any mutation by either method. We conclude that the PCR-SSCP analysis, performed as described here, is as sensitive and efficient as DNA sequencing in the ability to identify the sequence variations in the LDL receptor gene of the patients with heterozygous FH of this study.
Noncompetitive ELISA for human serum insulin-like growth factor-I
Authors: Khosravi M J , Diamandi A , Mistry J , Lee P D 
Published: January 2020

Abstract
Measurement of serum insulin-like growth factor I (IGF-I) is important in assessing the growth hormone/IGF axis. Competitive immunoassay methods for IGF-I are complicated by substantial interference from IGF-binding proteins (IGFBPs) due to the relatively low ratio of specific antibody to IGFBPs in the sample, even after standard acid-ethanol sample extraction. We report of development of a noncompetitive ELISA for human IGF-I that avoids this problem by utilizing excess amounts of capture and detection antibodies. Serum samples were prepared by using an abbreviated acid-ethanol extraction method. Neutralized supernatant was added to microwells coated with IGF-I capture antibody; horseradish peroxidase-labeled detection antibody was then added, incubated for 2 h, and then developed. Compared with the "gold standard" method of acid-column chromatography, the simplified acid-ethanol extraction yields a mean +/- SD recovery of 103% +/- 5.5% despite the presence of residual IGFBPs in the extracted sample. Comparisons with a centrifugal filtration sample extraction method are also shown. The ELISA is specific for IGF-I with an absolute sensitivity of 0.03 microgram/L and inter- and intraassay CVs of 3.9-8.8% and 2.6-6.7%, respectively. The availability of a rapid IGF-I ELISA combined with a simple and reliable sample preparation procedure should facilitate clinical and research studies of this important growth factor.
Effects of androgen suppression by gonadotropin-releasing hormone agonist and flutamide on lipid metabolism in men with prostate cancer: focus on lipoprotein(a)
Authors: Denti L , Pasolini G , Cortellini P , Ferretti S , Sanfelici L , Ablondi F , Valenti G 
Published: January 2020

Abstract
No clear relation between lipoprotein(a) [Lp(a)] and endogenous gonadal hormones has been demonstrated. In this study, we compared the effects on Lp(a) of pharmacological castration in 50 patients with prostate cancer who were undergoing therapy with a gonadotropin-releasing hormone agonist (goserelin), with effects on 58 age-matched controls. We also studied 16 untreated patients under baseline conditions and after 3 months of therapy with goserelin alone or combined with an antiandrogen (flutamide). Neither cross-sectional nor prospective studies showed any significant effects of therapy on Lp(a). However, cluster analysis identified a subgroup of patients showing slight but significant increases in Lp(a) concentrations, as well as greater declines of testosterone and estradiol, suggesting that androgen, like estrogen, can exert some slight, though not easily detectable, influence on Lp(a).
Distribution of blood viscosity values and biochemical correlates in healthy adults
Authors: Rosenson R S , McCormick A , Uretz E F 
Published: January 2020

Abstract
Increases in the viscosity of blood and plasma predict clinical manifestations of atherothrombotic vascular disease. The clinical utility of viscosity measurements in cardiovascular risk factor analysis requires reference values established from a healthy disease-free population. A cohort of 126 (71 men, 55 women) healthy nonsmoking adults had fasting blood analysis after a 12-14-h fast. Viscosity measurements were made on samples of whole blood, plasma, and serum at 37 degrees C with a coaxial cylinder microviscometer. The mean blood viscosity at shear rates of 100, 50, and 1 s-1 were 3.26 +/- 0.43, 4.37 +/- 0.60, and 5.46 +/- 0.84 mPa.s, respectively. Men had significantly higher blood viscosity values than women at each shear rate. The differences in blood viscosity did not remain significant after blood viscosity values were normalized to a hematocrit of 45%, except at 100 s-1. For the entire group, normalized blood viscosity values at each measured rate correlated inversely with HDL cholesterol and positively with fibrinogen. The mean plasma viscosity was 1.39 +/- 0.08 mPa.s and the mean serum viscosity was 1.27 +/- 0.06 mPa.s. Plasma viscosity correlated with fibrinogen (r = 0.51, P &lt; 0.0001), total serum protein (r = 0.33, P &lt; 0.0001), and triglyceride concentrations (r = 0.33, P &lt; 0.0015). Serum viscosity correlated with total serum protein (r = 0.50, P &lt; 0.0001) and LDL cholesterol (r = 0.24, P = 0.0065). This study provides reference values for the viscosity of blood, plasma, and serum that may assist in evaluating hemorheological profiles.
Evaluation of a two-site immunoradiometric assay for measuring noncomplexed (free) prostate-specific antigen
Authors: Cuny C , Pham L , Kramp W , Sharp T , Soriano T F 
Published: January 2020

Abstract
Serum prostate-specific antigen (PSA) in men is present as two different molecular forms separable by gel-filtration chromatography (GFC). We have evaluated a two-site IRMA that measures only the noncomplexed (free) form of PSA (F-PSA). Verification that the F-PSA assay measures solely F-PSA was obtained by assaying GFC-fractionated serum samples with both the F-PSA IRMA and a commercial PSA assay that measures total PSA (T-PSA: F-PSA plus alpha 1-antichymotrypsin-complexed PSA). The F-PSA assay detected only the 30-kDa peak corresponding to the free form of PSA, whereas the T-PSA assay detected two peaks: complexed PSA at approximately 90 kDa and F-PSA at approximately 30 kDa. The F-PSA assay had an analytical detection limit of 0.03 microgram/L and a measuring range up to 50 micrograms/L. The intraassay CV was 1.7-10% in the concentration range of 0.2-30 micrograms/L. The interassay CV was 3.4-12.5% in the same concentration range. Dilution and recovery studies showed no significant deviation from linearity across the assay range. The assay was insensitive to interference from hemoglobin, bilirubin, and total lipids up to concentrations of 5, 0.2, and 10 g/L, respectively. No significant loss of immunological activity (analyte stability) was seen day-to-day ( &lt; or = 5) or after repeated freeze/thaw ( &lt; or = 5) cycles. We conclude that the F-PSA IRMA is an accurate, precise, and reliable tool for measuring F-PSA in human serum.
Calpastatin autoantibodies: detection, epitope mapping, and development of a specific peptide ELISA
Authors: Schlosser U , Lackner K J , Scheckenhofer C , Schmitz G 
Published: January 2020

Abstract
Autoantibodies against calpastatin (CAST) were detected in a 53-year-old female patient with a history of arthritis and thrombosis. The specificity of the autoantibodies was determined by screening expression cDNA libraries, sequence analysis of positive clones, and subsequent Western blotting against recombinant antigen. Because the Western blot lacked satisfactory reproducibility, an ELISA for anti-CAST antibodies was established. The major epitope recognized by 24 Western blot-positive sera was located within the C-terminal 27 amino acids. The ELISA was therefore based on a synthetic peptide representing these amino acids. The assay was calibrated with serial dilutions of a positive reference serum. Intraassay precision is high with a CV of approximately 4% for low- and high-titer samples. Interassay precision (CV) was 5.6-8.2% for sera with low and intermediate titers [10-60 arbitrary units (AU), where 1:100 dilution of the positive reference serum = 100 AU, 1:1000 dilution = 10 AU, etc.], which increases with higher titers ( &gt; 60 AU). Among 205 healthy blood donors, the mean + 3 SD (after logarithmic transformation) was 30 AU; higher values were seen in 2.9% of 138 hospitalized patients. The newly developed ELISA will be a useful tool for further clinical studies on the association of anti-CAST antibodies to disease, because it permits rapid and reproducible analysis of patient sera.
Measurement of ascorbic acid in human plasma and serum: stability, intralaboratory repeatability, and interlaboratory reproducibility
Authors: Margolis S A , Duewer D L 
Published: January 2020

Abstract
We demonstrate that total ascorbic acid (TAA, the sum of ascorbic acid and dehydroascorbic acid) in properly prepared human plasma is stable at -70 degrees C for at least 6 years when preserved with dithiothreitol. TAA in human plasma or serum preserved with metaphosphoric acid degrades slowly, at the rate of no more than 1% per year. As assessed from our stability data and from data obtained from 23 laboratories over a period of &gt; 2 years, the intralaboratory repeatability of TAA measurement is approximately 2 mumol/L, irrespective of TAA concentration. Nonchromatographic analytical methods involving dinitrophenylhydrazine and 0-phenylenediamine yield biased results relative to chromatographic methods. Within groups of laboratories that use roughly similar analytical methods, the interlaboratory measurement reproducibility CV for TAA is 15%.
Differential diagnosis between hepatocellular carcinoma and cirrhosis through a discriminant function based on results for serum analytes
Authors: Castaldo G , Oriani G , Lofrano M M , Cimino L , Topa M , Budillon G , Salvatore F , Sacchetti L 
Published: January 2020

Abstract
We applied a multivariate analysis to a large series of serum biochemical tests in an attempt to identify a function that could efficiently discriminate cirrhosis from hepatocellular carcinoma (HC). We analyzed two successive temporal cohorts (1987-90; 1991-94) of HC and cirrhotic patients, all histologically classified (first cohort: 69 cirrhosis and 39 HC; second cohort: 66 cirrhosis and 38 HC). Using data from the first temporal cohort of patients, we obtained a discriminant function based on seven serum analytes: alpha-fetoprotein, the hepatic isoenzyme of alkaline phosphatase, lactate dehydrogenase isoenzyme 5, total gamma-glutamyltransferase (GGT), GGT isoforms complexed with low-density lipoprotein, aspartate aminotransferase, and copper. The same panel of analytes emerged when the second cohort was tested and also when both cohorts were tested together. In the two successive cohorts (total, 212 patients) with a prevalence of cirrhosis vs HC of approximately 2:1, the discriminant function correctly classified 93% of cases, the highest percentage of correct classification of the two diseases obtained so far by laboratory approaches. Validation with the jackknife reallocation statistical algorithm confirmed these results. In addition, of six patients with liver cirrhosis for whom we had the opportunity of following up and observing the evolution to HC, five were classified as HC at diagnosis by the multivariate discriminant analysis; i.e., discriminant analysis provided a diagnostic lead time of 6-12 months over histology. This discriminant function, based on easy-to-perform serum biochemical tests, may help solve a fundamental problem of differential diagnosis in the evolution of chronic liver diseases from cirrhosis to HC.
Quantification of vanadium in serum by electrothermal atomic absorption spectrometry
Authors: Heinemann G , Vogt W 
Published: January 2020

Abstract
We describe an ultrasensitive and reliable method for determining vanadium in human serum by electrothermal atomic absorption spectrometry. After lyophilization, the serum is digested in acid at high pressure, and the digests are evaporated to a small volume. Vanadium in the digests is complexed with cupferron, extracted, and dried. The residue is redissolved in formic acid, where it is 15-fold more concentrated than in the original serum sample. To enhance the furnace sensitivity, we injected six 40-microL aliquots (total, 240 microL) of the concentrated extract. The median concentration of vanadium in 108 persons was 50 ng/L, in good agreement with previously reported results by neutron activation analysis. The characteristic mass obtained (the mass required to give a signal of 0.0044A. s) was 28 pg, the limit of detection 11 ng/L, the limit of quantification 17 ng/L, and the total imprecision (CV) 5.5% at 1.54 micrograms/L. In two assays of Standard Reference Material 1577a (certified vanadium content 99 +/- 8 ng/g), we obtained values of 94.1 and 97 ng/g.
Epidemiology of illicit and abused drugs in the general population, emergency department drug-related episodes, and arrestees
Authors: Rouse B A 
Published: January 2020

Abstract
National trends in substance abuse are presented: the civilian noninstitutionalized general population; drug-related emergency department episodes; and booked arrestees. Major metropolitan differences are also noted. This study was based on the primary national data systems for these groups: The Substance Abuse and Mental Health Services Administration (SAMHSA) National Household Survey on Drug Abuse, SAMHSA's Drug Abuse Warning Network (DAWN), and the National Institute of Justice Drug Use Forecasting (DUF) system. While the most prevalent drug differed in the three data sources, all three showed recent increases in marijuana. Despite the general decline in drug use seen in the general population, both the number of drug-related cases in the DAWN system and the drug use detected in the DUF arrestees showed recent increases.
Preimplantation diagnosis by whole-genome amplification, PCR amplification, and solid-phase minisequencing of blastomere DNA
Authors: Paunio T , Reima I , Syvänen A C 
Published: January 2020

Abstract
We have developed a new method for preimplantation diagnosis of inherited diseases. Our procedure for the identification of point mutations in single cells combines whole-genome amplification using 15-mer random primers (primer extension preamplification, PEP) with a single locus-specific PCR amplification, followed by detection of the mutation by solid-phase minisequencing. The procedure was evaluated by detecting three disease-causing mutations and seven polymorphic nucleotides located on different human chromosomes from single granuloma and blastomere cells. The correct genotype of the cell was identified at 96% of the nucleotide positions analyzed, showing that a representative part of the genome is amplified during PEP. We estimate that PEP yielded at least 1000 copies of the genome. The quantitative nature of the solid-phase minisequencing method allowed us to notice that preferential amplification of one allele occurs at heterozygous loci during PEP, which is a potential problem in preimplantation diagnosis.
Analysis of low-dose benzodiazepines by HPLC with automated solid-phase extraction
Authors: Akerman K K , Jolkkonen J , Parviainen M , Penttilä I 
Published: January 2020

Abstract
We describe a simple, specific, and sensitive reversed-phase HPLC method with automated solid-phase extraction (SPE) for analyzing alprazolam, clonazepam, and nitrazepam concentrations in human serum or plasma. We prepare samples and calibrators with an automated sample preparer, using 100-mg Bond-Elut C18 SPE columns. The HPLC method uses isocratic elution with acetonitrile:methanol:dipotassium hydrogen phosphate, 10 nmol/L, pH 3.7 (30:2:100 by vol), at a flow rate of 1.5 mL/min to separate the drugs. We detect the benzodiazepines with diode-array detector at 240 nm; analyses of peak purity are performed at 210-365 nm. The recoveries were between 94% and 100%. The intraassay and interassay CVs were between 1.0% and 4.1%. The detection limit is 5 nmol/L. The antiepileptic and antidepressant drugs tested did not interfere with the assay. We developed the method for use in a clinical laboratory for therapeutic drug monitoring.
Sensitive and specific quantification of sirolimus (rapamycin) and its metabolites in blood of kidney graft recipients by HPLC/electrospray-mass spectrometry
Authors: Streit F , Christians U , Schiebel H M , Napoli K L , Ernst L , Linck A , Kahan B D , Sewing K F 
Published: January 2020

Abstract
Sirolimus (rapamycin) has a macrolide structure and is under clinical investigation as an immunosuppressant after organ transplantation. An HPLC/mass spectrometry assay to quantify sirolimus in blood was developed. 28-O-Acetyl sirolimus was used as internal standard. Blood samples were extracted with C18 columns. The extracts were injected into an HPLC system and isocratically eluted with methanol/1% formic acid (90/10 by vol) from a 150 X 4 mm C18 analytical column. The HPLC system was connected to a triple-stage quadrupole mass spectrometer with an electrospray interface and positive ions were detected. The limit of quantification in 1 mL of blood was 0.25 microgram/L and the calibration curve in blood was linear up to 250 microgram/L. The recovery from blood was 88 +/- 26% and interassay variation at 1 microgram/L was 19% and at 15 microgram/L 9.3%. Hydroxy, dihydroxy, demethyl, and didemethyl sirolimus as well as sirolimus were detected in blood of kidney graft patients.
Tacrolimus (FK506) metabolite patterns in blood from liver and kidney transplant patients
Authors: Gonschior A K , Christians U , Winkler M , Linck A , Baumann J , Sewing K F 
Published: January 2020

Abstract
The metabolite patterns of tacrolimus in blood were evaluated in 41 kidney and liver graft recipients. Trough concentrations of tacrolimus and its metabolites were measured by HPLC-mass spectrometry and microparticle enzyme immunoassay in parallel. A statistically significant correlation between results of both assays was observed for kidney and liver transplant patients (r = 0.77, P &lt;0.001 and r = 0.71, P &lt;0.001, respectively). The main metabolites in blood were demethyl, demethylhydroxy, didemethyl, didemethylhydroxy, and hydroxy tacrolimus. These metabolites added up to 42% (range 0-145%) of the tacrolimus concentration in liver transplant patients and to 44.8% (range 16-152%) in kidney transplant patients. During episodes of impaired liver function, concentrations of tacrolimus and its metabolites were increased compared with normal liver function, indicating accumulation of metabolites, in particular second-generation metabolites such as didemethyl and didemethylhydroxy tacrolimus. Stepwise regression analysis including tacrolimus, its metabolites, and liver function parameters suggested a model including serum activities of gamma-glutamyltransferase, alkaline phosphatase, and alanine aminotransferase as predictors for increased concentrations of demethyl tacrolimus, didemethyl tacrolimus, and the parent drug.