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June 1, 2005
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In a previous study which examined the distribution of apolipoprotein E genotypes and plasma levels in a sample of male coronary heart disease (CHD) patients and controls, we found a significant excess of the genotypes carrying APOE*4 allele in CHD men (18.2%) vs. controls (9.6%) and an association between the APOE*4 allele and the lowest concentrations of apoE. In the present investigation, we re-examined in the same samples two recently identified polymorphisms in the promoter region of APOE, −491A/T and −427T/C, which may alter the level of apoE expression. No differences in the distributions of the −491A/T genotypes and alleles were observed between cases and controls (−491*A = 0.760 and 0.757 respectively). Polymorphism −427T/C showed in CHD patients an excess of −427*C allele (patients vs. controls = 0.123 vs. 0.074) and corresponding genotypes that was marginally significant. Stratification of the samples according to the presence/absence of APOE*4 showed that the excess of the −427*C allele concerned only CHD patients not carrying APOE*4 allele (patients vs. controls = 0.133 vs. 0.061; p=0.017). This result suggests that the presence of −427*C allele could represent a risk for developing CHD in subjects with E2/E2, E3/E2, and E3/E3 genotypes. Studies carried out on patients with Alzheimer's disease demonstrated that −491A/T and −427T/C polymorphisms affect the level of plasma apoE. In the present study, carried out on CHD patients and controls, the genetic variation at −427 and −491 sites of the APOE regulatory region had no apparent effect on apoE plasma concentration.
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June 1, 2005
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A constitutive isotype of human serum amyloid A, serum amyloid A4 (SAA4), is distributed into plasma lipoproteins, primarily in high density lipoproteins. Its physiological function is unknown; its serum concentration has no relationship with those of other major apolipoproteins. In this study, changes in SAA4 concentrations were further characterized. Variations in healthy individuals were negligible. In subjects under-going renal allograft transplantation, SAA4 changed in parallel with acute phase SAA, although its magnitude was not larger than a three-fold increase. This confirmed that SAA4 is a minor acute phase reactant in humans. SAA4 concentrations showed a good agreement with serum pseudocholinesterase activity in healthy subjects and patients with lowered pseudocholinesterase when patients with elevated acute phase SAA were excluded. These results suggest that SAA4 can be an indicator of nutrition or of hepatic protein synthesis in the absence of inflammation.
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June 1, 2005
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The D polymorphism of angiotensin converting enzyme (ACE) gene has been found to be associated with various diseases, and ACE may also be involved in the pathogenesis of erectile dysfunction. On the other hand, interpretation of the data on the association of DD genotype with various diseases is controversial, due to methodological and technical variations in detection of the polymorphisms. We investigated a possible association between the DD genotype and erectile dysfunction in a Korean population, and compared the frequency of ACE genotypes using our multiplexed PCR method with those based on the conventional PCR method in a sample of erectile dysfunctional and control subjects. There was significant difference in the distribution of ACE genotypes between the erectile dysfunctional (conventional PCR) and the control subjects (multiplexed PCR) (χ 2 =7.395, p<0.05), but there was no significant difference in the distribution of the genotypes between both groups (χ 2 =0.815, p<0.05) when our multiplexed PCR method was used. Therefore our results suggest that especially the conventional PCR method for ACE gene polymorphism may require careful control and may need repeated testing to verify the insertion deletion (ID) heterozygotes, and that a multiplexed PCR method can markedly increase the detection rate of the I allele in ID heterozygotes. No association was found between I/D polymorphism and erectile dysfunctional subjects in the Korean population studied.
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June 1, 2005
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In spite of the fact that pouchitis is the most frequently occurring and troublesome complication found in patients treated by ileo-anal anastomosis for ulcerative colitis, no biological marker currently exists to monitor the outcome of the disease. Since it has been noted faecal butyrate is reduced in patients with pouchitis, we developed a simple gas chromatography method to quantify butyrate in faecal water. This test is based on diethyl ether extraction with the use of methacrylic acid as an internal standard. We demonstrated that butyrate was effectively measured when this technique was applied to eleven patients with ileal-pouch anal anastomosis within the first year after the closure of their ileostomy. We also observed a noticeable reduction in the concentration of butyrate in patients who went on to develop a pouchitis.
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June 1, 2005
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The plasma concentration of glycerol, the backbone of triglycerides and the end product of triacylglycerol breakdown, is considered to reflect lipolysis in adipose tissue. We evaluated an automated enzymatic procedure for the measurement of glycerol in plasma. The assay was linear up to 250μmol/l. The detection limit was 8μmol/l. Recovery averaged 94% from spiked plasma and 104% from diluted plasma. An extensive precision study performed according to the guidelines of the National Committee for Clinical Laboratory Standards showed within-run coefficients of variation between 14.8% and 2.6% for concentrations ranging from 28μmol/l to 164 μmol/l. The reference range for fasting healthy adult men was 14–69 μmol/l. Glycerol levels were significantly correlated with free fatty acid levels. This automated enzymatic method is rapid and reliable, and provides greater sensitivity or convenience than previously described procedures.
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June 1, 2005
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Detection of autoantibodies to the thyrotropin receptor by radioreceptor assays is largely requested in clinical practice for the diagnosis of Graves' disease and its differentiation from diffuse thyroid autonomy. Additionally, thyrotropin receptor antibodies (TRAb) measurement during antithyroid drug treatment can be useful to evaluate the risk of relapse after discontinuation of the therapy. Nevertheless, some patients affected by Graves' disease are TRAb-negative when a 1st generation assay is used. In this study we evaluated the diagnostic performance of a newly developed 2nd generation TRAb assay in 46 patients with Graves' disease with negative 1st generation TRAb assay results. A control group of 50 Graves' disease patients with positive 1st generation TRAb assay results, 50 patients with Hashimoto's thyroiditis and 50 patients with nodular goiter were also examined. Forty one of 46 patients with Graves' disease with negative 1st generation TRAb assay results showed a positive 2nd generation test. No differences were seen in control groups. In conclusion, the 2nd generation TRAb assay is more sensitive than the 1st generation test and should be used in clinical practice. Long-term prospective studies are needed to evaluate the prognostic role of the 2nd generation TRAb assay in Graves' disease.
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June 1, 2005
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The aim of this study is to determine whether the presence of microtransferrinuria and microalbuminuria detected in pregnant women who are free of symptoms can predict the subsequent development of preeclampsia. One hundred fifty five pregnant women were successfully followed from 10 weeks gestation up till delivery. Pre-eclampsia developed in 31 women (17 mild and 12 severe pre-eclampsia), and eclampsia developed in two cases, whereas 124 women remained normotensive (controls). First morning urine specimens were collected during 10 to 12 weeks gestation and analyzed for microalbuminuria by a specific immunochemical test strip method. Mid-trimester mean a rterial blood pressure (MAP) was also measured. Urinary microtransferrin levels in pregnant women who subsequently developed severe pre-eclampsia and eclampsia were significantly higher than those of pregnant women who remained normotensive. Microtransferrinuria as a predictor for pre-eclampsia had a sensitivity 93.5%, specificity 65%, positive predictive value 83% and negative predictive value 98.4%, whereas these values for microalbuminuria were: 50%, 58%, 50% and 91%, respectively. Urinary microtransferrin levels were significantly elevated in women with elevated MAP and in women who delivered low birt h weight and low Apgar score babies. In conclusion, microtransferrinuria is a potentially more sensitive predictor of pre-eclampsia than microalbuminuria. Moreo v e r, microtransferrinuria in early pregnancy might be a negative marker of fetal outcome in pre-eclampsia.
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June 1, 2005
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Cholecystectomized patients with recurrent attacks of right epigastric pain and with dilated common bile duct are a clinical challenge. In a number of these patients dilatation of the common bile duct is explained as a normal postoperative state following cholecystectomy and the recurrent pain attacks are of origin other than bile disorder, but in some cases dilatation of the common bile duct and attacks are caused by bile duct stones. The aim of the present work was to study the value of common plasma liver function tests in predicting bile duct stones in the group of non-icteric cholecystectomized patients with recurrent attacks of right epigastric pain and with dilated common bile duct. The study population comprised 24 consecutive non-icteric cholecystectomized patients admitted for elective endoscopic retrograde cholangiopancreatography because of attacks of right epigastric pain and dilated common bile duct in ultrasonography. All the liver function tests seemed to assist in separating patients with bile duct stones (n=11) from those without (n=13). Alanine aminotransferase levels were significantly higher (p=0.05) in patients with bile duct stones than in those without, but also alkaline phosphatase (p=0.07), γ-glutamyl transferase (p=0.09) and bilirubin (p=0.09) levels seemed to be higher in patients with bile duct stones than in those without, although the differences in these values did not reach statistical significance. In conclusion, common plasma liver function tests assist in separating patients with bile duct stones from those without in this small but clinically important group of non-icteric cholecystectomized patients with recurrent attacks of right epigastric pain and with dilated common bile duct. However, the actual value of these measurements is limited in clinical decision making since overlapping of values occured.
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June 1, 2005
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The International Germ Cell Cancer Collaborative Group study of patients with metastatic testicular germ cell tumors showed that catalytic concentration of serum lactate dehydrogenase (S-LD), serum α-fetoprotein concentration (S-AFP), and serum human chorionic gonadotropin concentration (S-hCG) predicted death from tumor. The recent international TNM classification (T primary tumor, N lymph node metastasis, M distant metastasis) is based on these results. The aim of our study was to evaluate whether catalytic concentration of S-LD isoenzyme 1 (S-LD-1) was a better predictor than the criteria used for the international classification. In an evaluation series of 44 patients from Odense University Hospital, Denmark, a raised S-LD-1 (>1.0 x upper limit of reference values) had a predictive value for death from tumor in 5-years observation of 46%. The predictive value was 46% for S-LD, 25% for S-AFP, and 40% for S-hCG. A normal SLD-1 had a predictive value for survival over 5-years observation of 100%. It was 81% for S-LD, 75% for S-AFP, and 77% for S-hCG. The fraction of the patients who died of tumor and had a raised tumor marker value was 100% for S-LD-1, 46% for S-LD, 9% for S-AFP, and 18% for S-hCG. The fraction of patients with a normal serum tumor marker value among those who survived was 61% for S-LD-1, 81% for S-LD, 94% for S-AFP, and 94% for S-hCG. A validation series of 37 patients treated at the University of Texas MD Anderson Cancer Center showed similar findings. Combining the patients in the two series, a raised value of SLD-1 classified more patients into a subgroup with an impaired survival (53%) than S-LD (35%), S-AFP (6%), or S-hCG (11%), and the high risk subgroups based on the international classification (40%). The findings have implications for the staging and treatment of patients with metastatic testicular germ cell tumors.
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June 1, 2005
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The annual inter- and intra-individual biological variation, including the circannual rhythmic variation, of the serum concentrations of magnesium and ionized calcium has been investigated in a group of 51 apparently healthy volunteers. Venous blood specimens were collected on intervals of once a month within a one-year period, using a standardized protocol. The inter-individual coefficients of variation were 5.12% for magnesium and 1.58% for ionized calcium. The medians of the intra-individual coefficients of variation were 1.93% for magnesium and 2.18% for ionized calcium. These data were used to determine the allowable imprecision, the allowable systematic error, the critical difference for significant change detection, and the usefulness of population reference values (index of individuality). Of the quantities studied, only the serum concentration of ionized calcium shows a significant annual rhythmic variation (amplitude 1.3%), although this result may be due to the between-run metrological variance, considering that the concentration of ionized calcium of the control material used during the study possesses a similar significant rhythmic variation.
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June 1, 2005
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A variety of laboratory measurement units, assay systems, and reference values are presently being used in different clinical environments. There is therefore a challenge in provision of tools for presentation of laboratory data in harmonized forms that reduce the information load and reinforce clinical perception of test results. This paper describes the use of standard deviation (SD) units and logarithmic time graphical displays to improve presentation of laboratory test results. The SD concept employs strength of the change from normality as a diagnostic indicator, and the logarithmic time scale enables long-term overview of patient results. The suggested format is expected to promote effective transfer of test result information between laboratories, clinicians and hospitals.
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June 1, 2005
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Statistical software often offers a list of various descriptive statistics of location and scale, but rarely selects an efficient estimate that is statistically adequate for an actual univariate sample. The sample interval estimate for a specified degree of uncertainty seems to be more meaningful if it covers an unknown value of the population parameter. The concept of an interval estimate in medicine is then used for medical decision-making. The proposed methodology, which uses the S-Plus algorithm for biochemical, biological and clinical data analysis contains the following steps: (i) Exploratory data analysis identifies basic statistical features and patterns of the data, the distributions of which are mostly non-normal, non-homogeneous and often corrupted by outliers. (ii) Sample assumptions about data, independence of sample elements, normality and homogeneity are examined. (iii) Power transformation and the Box-Cox transformation to improve sample symmetry and stabilize the spread. (iv) Classical and robust statistics for both large ( n >30) and medium-sized samples (15< n <30), point and interval estimates for the parameters of location, scale and shape. For an analysis of small samples (4 < n < 20) the Horn procedure of pivot measures is recommended. The proposed methodology is demonstrated in two case studies, a large sample analysis of mean pregnenolone concentrations in the umbilical blood of newborns, and a small sample analysis of mean haptoglobin concentrations in human serum.
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June 1, 2005
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In the recent years the number of commercially available immunoassays for the detection of human cytomegalovirus (HCMV)–specific immunoglobulin M (IgM) antibodies has rapidly increased. The aim of the present study was to evaluate five commercial immunoassays for the serological diagnosis of HCMV-infection. These methods, namely the IMx CMV IgM assay, the AxSYM CMV IgM assay (both Abbott), the Gull CMV IgM, the CMV-IgM-ELA test PCS Medac and the Biotest Anti-HCMV recombinant IgM ELISA, were compared for their diagnostic effectiveness and interference with substances eventually producing cross-reactions with HCMV-IgM (Epstein-Barr-virus (EBV)-IgM, rheumatoid factor (RF)). In addition, repeated measurements on samples from kidney and heart transplant recipients with active HCMV infection were examined to compare the temporal development of the HCMV-IgM measured with the five assay systems. Since there is no commercially available gold standard, it was assumed that the true classification, of whether the patient sample is HCMV-IgM positive or negative, was unknown. Hence sensitivity and specificity were assessed based on a maximum likelihood approach using a “latent class” model. The cross-reactions were quantified by a Bayesian statistical model using prior information for the expected prevalences in the EBV-IgM and rheumatoid factor sample groups. The results of the study demonstrated that there are great differences in sensitivity and specificity as well as in cross-reactions with EBV-IgM and RF between the tested ELISAs.
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June 1, 2005
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A new reagent carrier, Reflotron ® ALP, has been developed for the Reflotron ® system, allowing easy and rapid measurement (in less than 3 minutes) of alkaline phosphatase (ALP) activity in capillary blood, venous blood, heparinized plasma or serum. The evaluation of the analytical performance of the assay was carried out at eight clinical laboratories. The study of the imprecision using the measurements in human samples resulted in coefficients of variation ranging from 1.3% to 4.6% (within-run) and from 3.2% to 4.0% (day-to-day). The analytical specificity of the Reflotron ® ALP assay agrees well with ALP methods using a N-methyl-D-glucamine buffer solution. The calibration of the Reflotron ® ALP assay, however, is related to the reference intervals for ALP methods using a diethanolamine buffer solution. Method comparisons were performed with the ALP method on Hitachi instruments using diethanolamine buffer. Reflotron ® ALP measurements in blood and plasma in 157 randomly selected split samples showed excellent agreement (slope: 0.99; intercept: 0.7 U/l; median bias: 2.3%; median difference from the comparison method: −0.3%). Specimens from pregnant women and adolescents were excluded from this study. Differing values were obtained in a method comparison using 48 samples containing predominantly the ALP bone isoform (slope: 0.81; intercept: 31.5 U/l; median bias: 5.7%; median difference from the comparison method: −12.2%). Regression analysis of the results from 21 sera with prevailing placental ALP gave a slope of 1.51, and an intercept of −41.1 U/l (median bias: 8.6%; median difference from the comparison method: 35.6%). Reflotron ® ALP was compared with three different wet chemistry procedures using different buffer compounds: N-methyl-D-glucamine or diethanolamine or 2-amino-2-methyl-1-propanol. In samples containing predominantly ALP isoforms not of liver origin, the measurements with N-methyl-D-glucamine buffer gave the best fit with respect to Reflotron ® . In an interference study with 18 drugs, no effect on the test results could be detected. Total bilirubin up to 750 μmol/l and hemolysis up to 1.7 g/l free hemoglobin did not influence the test. Reflotron ® ALP proved to be an easy and rapid method with excellent precision. The accuracy related to an ALP method using diethanolamine buffer was good. The systematic differences for ALP in samples from pregnant women and adolescents have to be taken into account. The assay is well suited for differential diagnosis of hepatic diseases in decentralized testing.
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July 27, 2005
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