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Selected AbstractsDiagnosis of Chlamydia trachomatis infectionJOURNAL OF CLINICAL LABORATORY ANALYSIS, Issue 1 2006Jayanti Mania-Pramanik Abstract Important progress in the diagnosis of Chlamydia trachomatis (C. trachomatis) includes the development of nucleic acid amplification techniques such as polymerase chain reaction (PCR) and ligase chain reaction (LCR). Commercial kits are available, but they are costly, sporadic in availability, must be imported, and are economically beyond the reach of common people. To overcome this limitation, most research laboratories have standardized their in-house-developed PCR methods for diagnosing this infection. However, each laboratory has to spend a great deal of time and money to accomplish this. Published reports do not always elaborate the steps involved in standardizing a test so that it can immediately be reproduced in another setting. In the present study we attempted to elaborate the steps involved in standardizing a sensitive and specific PCR technique followed by hybridization with specific C. trachomatis probe to diagnose this infection in cervical, introital, and urine specimens, and used it to determine the infection rate in a clinical population. J. Clin. Lab. Anal. 20:8,14, 2006. © 2006 Wiley-Liss, Inc. [source] Polymorphisms of Alcohol Metabolizing Enzymes in Indigenous Mexican Population: Unusual High Frequency of CYP2E1*c2 AlleleALCOHOLISM, Issue 1 2010Elizabeth Gordillo-Bastidas Background:, Alcohol abuse represents the major identified etiological factor of cirrhosis in México. ADH1B, ALDH2, and CYP2E1 have been considered candidate genes in alcohol-related diseases. Controversial results probably due to ethnic differences, among other factors, have been reported. Mexican Mestizos (MES) derive from the combination of indigenous, Spaniard, and African genes. Huichols (HUI) constitute an indigenous group from western Mexico with no racial admixture. We determined ADH1B*2, ALDH2*2, and CYP2E1*c2 allele frequencies in healthy HUI and MES from western Mexico. Lipid and hepatic profile were also carried out. Methods:, One hundred and one HUI and 331 MES subjects were studied. Genotype and allele frequency were assessed through polymerase chain reaction,restriction fragment length polymorphism after DNA isolation from peripheral leukocytes. Commercial kits for lipid and hepatic determinations were used. Results:, Polymorphic allele distribution in HUI was: 0%ADH1B*2, 0.5%ALDH2*2, 51.5%CYP2E1*c2; in MES: 3.4%ADH1B*2, 0%ALDH2*2, 16.1%CYP2E1*c2. Frequency of ADH1B*2 was statistically (p < 0.001) lower in HUI than MES. CYP2E1*c2 polymorphic allele was significantly higher (p < 0.0001) in HUI than MES. Hepatic profile was normal in both groups. HUI showed a better lipid profile than MES independently of genotype. Conclusions:, Huichols exhibited the highest CYP2E1*c2 allele frequency of the world documented up to this date; meanwhile, ADH1B*2 and ALDH2*2 were practically absent. This feature could be useful in the understanding of Mexican population gene composition, alcohol metabolism, and alcoholic liver disease development. However, further association studies are necessary. The heterogeneity of Mexican population was evidenced by the significantly different distribution of CYP2E1*c2 allele observed among different regions of the country. Lipid and hepatic values were not associated to genotype. This report constitutes the first study dealing with gene polymorphisms of alcohol metabolizing enzymes conducted in HUI. [source] "Paterniplex", a highly discriminative decaplex STR multiplex tailored for investigating special problems in paternity testingELECTROPHORESIS, Issue 21 2007Thomas Betz Abstract The goal of the study was to develop a STR multiplex ("Paterniplex") that is , as supplement to commercially available multiplex kits like the Identifiler® kit (Applied Biosystems, Foster City, CA) , suitable for solving complex paternity cases such as deficiency cases or cases with mutations. The Paterniplex comprises the nine highly polymorphic STRs D8S1132, D7S1517, D10S2325, D12S391, Se33, D17S976, Penta E, Penta D and FGA in addition to Amelogenin as sex determination marker. The loci were selected because of their high degree of polymorphism (higher than that of the widely used TH01 marker). Only one locus, FGA, is shared with the Identifiler kit to avoid sample mix up. The study further gives details on the population genetics of the loci in a German Caucasian population (allelic distribution, Hardy,Weinberg Equilibrium and forensic efficiency markers such as the Discriminating Power) and three examples for cases that could not be solved using commercially available kits alone, but using the Paterniplex in addition to a commercial kit. [source] Automatic analysis of multiplex ligation-dependent probe amplification products (exemplified by a commercial kit for prenatal aneuploidy detection)ELECTROPHORESIS, Issue 22 2005Tommy Gerdes Dr. Abstract For use in routine prenatal diagnostics, we developed software and methods for automatic aneuploidy detection based on a commercial multiplex ligation-dependent probe amplification (MLPA) kit. Software and methods ensure a reliable, objective, and fast workflow, and may be applied to other types of MLPA kits. Following CE of MLPA amplification products, the software automatically identified the peak area for each probe, normalized it in relation to the neighboring peak areas of the test sample, computed the ratio relative to a reference created from normal samples, and compensated the ratio for a side effect of the normalization procedure that scaled all chromosomally normal DNA peak areas slightly up or down depending on the kind of aneuploidy present. For the chromosomes 13, 18, 21, X, and Y, probe reliability weighted mean ratio values and corresponding SDs were calculated, and the significance for being outside a reference interval around ratio 1.0 was tested. p,,,1% suggested aneuploidy and 1,<,p,,,5% suggested potential aneuploidy. Individual peaks, where the normalized area was situated more than 4 SD from the corresponding reference, suggested possible partial deletion or gain. Sample quality was automatically assessed. Control probes were not required. Having used the software and methods for two years, we conclude that a reliable, objective, and fast workflow is obtained. [source] DNA extraction procedure: a critical issue for bacterial diversity assessment in marine sedimentsENVIRONMENTAL MICROBIOLOGY, Issue 2 2006Gian Marco Luna Summary In order to evaluate whether different DNA extraction procedures can affect estimates of benthic bacterial diversity, based on 16S rRNA gene terminal restriction fragment length polymorphism (T-RFLP) fingerprinting technique, we compared two in situ lysis procedures (a SDS-based protocol and a commercial kit for DNA recovery) and one cell-extraction protocol on a variety of marine sediments. Despite the two in situ lysis procedures resulted in significantly different DNA yields (highest with the SDS in situ lysis), estimates of bacterial diversity provided a not significantly different ribotype richness, as well as similar values of the Shannon-Wiener (H,) and Margalef (d) indices of biodiversity and of evenness (Pielou index, J). Conversely, the cell-extraction procedure for DNA extraction resulted always in a significantly lower ribotype richness and diversity. The analysis of similarities (anosim) among the T-RFLP electropherograms allowed concluding that ribotypes composition did not change significantly using different protocols. However, the analysis of ,-diversity (turnover diversity) revealed that a large number of ribotypes was observed exclusively with one of the three protocols utilized. When unshared ribotypes from in situ lysis and cell extraction were pooled together, total ribotype richness resulted much higher (up to 80%). Our results indicate that estimates of ribotype diversity based on a single protocol of DNA extraction can significantly underestimate the total number of bacterial ribotypes present in the benthic domain. We recommend that future studies will not only integrate different DNA extraction procedures, but also will explore the possibility of integrating two or more different genetic markers in order to increase our ability to detect the actual bacterial diversity in environmental samples. [source] A novel role of alkaline phosphatase in protection from immunological liver injury in miceLIVER INTERNATIONAL, Issue 1 2002Qiang Xu Abstract:Aims/Background: Little is known about the role of alkaline phosphatase (AP) in liver diseases, except for its elevation in jaundice or cholestasis. Its substrate, endotoxin, is usually elevated in patients as well as animals with liver damage. This study aimed to provide evidence for its new role as protection against immunological liver damage. Methods: Liver injury was induced in mice by delayed-type hypersensitivity to picryl chloride. AP activity was measured using a commercial kit. Results: In acute liver injury, a significant decrease in AP activity in serum was observed but there was an increase in liver tissue. Single administration of cyclophosphamide before sensitization with picryl chloride exacerbated the liver injury, with more serious AP changes, while consecutive use after the sensitization alleviated the injury with a recovery from the changes. When liver injury proceeded for 1 week, both serum and liver showed decreased AP activity. Lipopolysaccharide facilitated alanine transaminase release from levamisole-pretreated but not non-treated hepatocytes from naive mice. However, the release was confirmed from liver slices of mice with liver injury proceeding for 1 week, even without levamisole pretreatment. Conclusion: The development of liver injury may lead to a dysfunction in AP synthesis and release. Levamisole may make normal hepatocytes, like the hepatocytes from liver-injured mice, highly sensitive to lipopolysaccharide through inhibiting AP synthesis. The findings obtained in this study suggest that AP may contribute to protection from injury by a mechanism involving neutralization of endotoxin. [source] Development of recombinant OmpA and OmpB proteins as diagnostic antigens for rickettsial diseaseMICROBIOLOGY AND IMMUNOLOGY, Issue 7 2009Eun-Ju Do ABSTRACT In this study the diagnostic potential of Rickettsia conorii recombinant antigens was analyzed. For this, site-specific PCR primers were used to clone the OmpA and OmpB genes of R. conorii into pMAL-c2X plasmids. Six fragments of OmpA and four of OmpB were expressed as fusion proteins with maltose-binding protein in Escherichia coli. OmpA1350-1784, OmpB801-1269, and OmpB1227-1634 regions from truncated proteins were selected as diagnostic candidate antigens by ELISA using control sera. ELISA results of three antigens were compared to the results obtained by using a commercial ELISA kit which contained whole OmpA and OmpB antigens from R. conorii. For this analysis, 40 serum samples taken from febrile patients and uninfected controls were tested. Of the 20 R. conorii test results which were positive with the commercial kit, 18 were shown to be positive by ELISA using OmpA1350-1784 (a sensitivity of 90%). The specificity of the ELISA was 100%; all of the 20 samples shown to be negative using the commercial kit were also negative in our assay. The sensitivities of the ELISA using the OmpB801-1269 and OmpB1227-1634 were 90% and 95%, respectively. The specificities of the OmpB801-1269 and the OmpB1227-1634 were 100% and 95%, respectively. These results suggest that specific regions of OmpA and OmpB effectively detect antibodies against R. conorii, and the truncated recombinant antigens could be used for development of diagnostic tools for rickettsial disease. [source] Circulating tumour-associated plasma DNA represents an independent and informative predictor of prostate cancerBJU INTERNATIONAL, Issue 3 2006FELIX K.-H. OBJECTIVE To investigate whether preoperative plasma levels of free DNA can discriminate between men with localized prostate cancer and benign prostatic hyperplasia (BPH). PATIENTS AND METHODS In all, 161 referred patients suspicious for prostate cancer either by an elevated prostate-specific antigen (PSA) level and/or abnormal digital rectal examination (DRE) were included in this prospective study. Peripheral plasma was taken before prostate biopsy and genomic DNA was extracted from the plasma using the a commercial kit and a vacuum chamber. After controlling for age, PSA level, the percentage free/total (f/t) PSA and prostate volume, the median prostate cancer plasma DNA concentration served as diagnostic threshold in uni- and multivariate logistic regression models. Multivariate models were subjected to 200 bootstraps for internal validation and to reduce over-fit bias. RESULTS Subgroups consisted of 142 men with clinically localized prostate cancer and 19 with BPH. The median plasma concentration of cell-free DNA was 267 ng/mL in men with BPH vs 709 ng/mL in men with prostate cancer. In univariate analyses, plasma DNA concentration was a statistically significant and informative predictor (P = 0.032 and predictive accuracy 0.643). In multivariate analyses, it remained statistically significant after controlling for age, tPSA, f/tPSA and prostate volume, increasing the predictive accuracy by 5.6%. CONCLUSIONS Our data suggest that plasma DNA level is a highly accurate and informative predictor in uni- and multivariate models for the presence of prostate cancer on needle biopsy. The predictive accuracy was substantially increased by adding plasma DNA level. However, larger-scale studies are needed to further confirm its clinical impact on prostate cancer detection. [source] Antidiabetic and toxicological evaluations of naringenin in normoglycaemic and NIDDM rat models and its implications on extra-pancreatic glucose regulationDIABETES OBESITY & METABOLISM, Issue 11 2008R. R Ortiz-Andrade Aim:, The present investigation was designed to determine the in vivo antidiabetic effect of naringenin (NG) in normoglycaemic and diabetic rat models through blood glucose (GLU) measurements following acute and subchronic time periods. Possible modes of action of NG were investigated and its acute toxicity determined. Methods:, Normoglycaemic and non-insulin-dependent diabetes mellitus (NIDDM) rat models were treated for acute and subchronic (5 days) time periods with 50 mg/kg/day of NG. Blood biochemical profiles were determined after 5 days of the treatment in normoglycaemic and NIDDM rats using commercial kits for GLU, triglycerides (TG), total cholesterol (CHOL) and high-density lipoprotein (HDL). In order to elucidate its antidiabetic mode of action, NG was administered intragastrically and an oral glucose tolerance test performed using GLU and sucrose (2 g/kg) as substrates. The inhibitory effect of a single concentration of NG (10 ,M) on 11,-hydroxysteroid dehydrogenase type 1 (11,-HSD1) activity in vitro was determined. Finally, the preclinical safety and tolerability of NG was determined by toxicological evaluation in mice and rats using Organization for Economic Cooperation and Development (OECD) protocols. Results:, Intragastrically administered NG (50 mg/kg) induced a significant decrease in plasma GLU in normoglycaemic and NIDDM rat models (p < 0.05) following acute and subchronic time periods. After 5 days of administration, NG produced significant diminished blood GLU and TG levels in streptozotocin,nicotinamide,induced diabetic rats. The administration of NG to normal rats significantly increased the levels of TG, CHOL and HDL (p < 0.05). NG (5 and 50 mg/kg) induced a total suppression in the increase of plasma GLU levels after administration of substrates (p < 0.01), but NG did not produce inhibition of ,-glucosidase activity in vitro. However, NG (10 ,M) was shown to inhibit 11,-HSD1 activity by 39.49% in a cellular enzyme assay. Finally, NG showed a Medium Lethal Dose LD50 > 5000 mg/kg and ranking at level five based on OECD protocols. Conclusion:, Our findings suggest that NG may exert its antidiabetic effect by extra-pancreatic action and by suppressing carbohydrate absorption from intestine, thereby reducing the postprandial increase in blood GLU levels. [source] CSF biomarker profile and diagnostic value in vascular dementiaEUROPEAN JOURNAL OF NEUROLOGY, Issue 2 2009G. P. Paraskevas Background and purpose:, The differential diagnosis between vascular dementia (VD) and Alzheimer's disease (AD) or mixed dementia (MD) is not always easy in clinical practice. The purpose of the present study was to evaluate the cerebrospinal fluid (CSF) biomarkers tau protein in its total (,T) or hyperphosphorylated at threonin-181(,P-181) form and beta amyloid peptide 1,42 (A,42) alone and their combinations to investigate their diagnostic value in the discrimination between VD and AD or MD. Methods:, The above CSF biomarkers were determined in duplicate and blind to the clinical diagnosis by double sandwich, enzyme-linked immunosorbent assay (ELISA) commercial kits (Innogenetics, Gent, Belgium) in 92 AD patients, 23 VD patients, 17 patients with MD and 68 controls. Results:, Alzheimer's disease and MD showed increased levels of ,T, ,P and reduced levels of A,42 as compared with the controls. The best discrimination between VD and AD or MD was achieved by the combination of all three biomarkers, correctly classifying ,85% of patients, either in the form of a discriminant function or in the form of the ,T × ,P-181/A,42 formula. Conclusions:, Cerebrospinal fluid biomarkers may be a useful adjunct for the discrimination between AD/ MD and VD in every day clinical practice. [source] Potentiation of isoniazid-induced liver toxicity by rifampicin in a combinational therapy of antitubercular drugs (rifampicin, isoniazid and pyrazinamide) in Wistar rats: A toxicity profile studyHEPATOLOGY RESEARCH, Issue 10 2007Sheikh Abdullah Tasduq Aim:, Biochemical characterization of long-term toxic manifestations of anti-tubercular (anti-TB) drugs , rifampicin (RIF), isoniazid (INH) and pyrazinamide (PZA) , individually and in two combinations: (i) RIF + INH, and (ii) RIF + INH + PZA in Wistar rats. Methods:, Animals received anti-TB drugs , alone or in combination , once daily p.o. for up to 90 days (doses, in mg/kg: RIF, 250; INH, 50; PZA, 100). Assays for alanine aminotransferase (ALT), alkaline phosphatase (ALP), bilirubin (serum) and lipid peroxidation (LPO), glutathione (GSH), glutathione peroxidase (GPx), catalase, Na+K+-ATPase and CYP 2E1 (liver) were performed to assess liver toxicity. Clinical biochemistry was done by commercial kits. Determinations were made at 0, 15, 30 and 90 days of treatment schedule. Results:, Anti-TB drugs-treated animals showed abnormal rises or falls (>1.5,2 fold) in the serum/liver parameters. Mild hyperlipidemia, hypercholesterolemia and hyperuricemia were the other pathologies. Of all the treated groups, INHalone or in combination with other drugs produced a progressive enhancement of toxicity over 15,90 days. The in vivo results were further supported by in vitro results (MTT assay, GSH and LPO) in primary cultures of rat hepatocyte. Results indicated that anti-TB drugs in combination: (i) caused membrane damage resulting in leakage of ALT, ALP and bilirubin; (ii) caused imbalance in endogenous enzymatic oxidant,antioxidant defense via increased lipid peroxidation and in glutathione homeostasis; and (iii) enhanced the CYP 2E1-mediated bioactivation mechanism. Conclusion:, Toxicity manifestations seemed to be heptocytic injury targeted at hepatocytes, bile ducts or sinusoidal cells related to hepatitis and primary biliary cholestasis. [source] Age-related plasma reference ranges for two heparin-binding proteins , vitronectin and platelet factor 4INTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY, Issue 6 2009F. NEWALL Summary This study was conducted to establish age-related reference ranges for two heparin-binding proteins , vitronectin and platelet factor 4 (PF4) , and to determine if the quantitative values of these proteins may contribute to the reported age-dependent effect of unfractionated heparin (UFH). Plasma samples were obtained from healthy children aged between 1 month and 16 years and from healthy adult volunteers. Two commercial kits were used to measure plasma vitronectin and PF4 levels. Results were reported as mean and boundaries including 95% of the population. Plasma vitronectin levels for children aged 1,5 years were significantly higher compared with adults. Plasma PF4 levels for infants <1 year of age were significantly lower compared with adults. The differences between reference values for both proteins in all other age-groups were not statistically significant. This study for the first time has established age-related reference ranges for vitronectin and PF4. In establishing these ranges, the quantitative values of these proteins do not appear to be the major contributory cause for the age-dependent variation in UFH effect. Future studies are required to evaluate the possible impact of age-dependent differences in binding between heparin-binding proteins and UFH. [source] Role of anti-transglutaminase (anti-tTG), anti-gliadin, and anti-endomysium serum antibodies in diagnosing celiac disease: A comparison of four different commercial kits for anti-tTG determinationJOURNAL OF CLINICAL LABORATORY ANALYSIS, Issue 3 2001D. Basso Abstract The aims of this study were: (1) to compare the diagnostic efficacy for celiac disease (CD) diagnosis of serum determination of anti-gliadin (AG) (IgA and IgG) and anti-endomysium (AE) with that of anti-transglutaminase (AtTG); and (2) to compare the accuracy of four different assays to measure AtTG. We studied 72 children: the histological diagnosis of CD was made in 38 cases and excluded in the remaining 34 children. In fasting sera we measured AE, AG-IgA and IgG, and AtTG, the latter with four different commercial kits (Eurospital, Medipan, Inova, Arnika). Moreover AtTG was measured in a group of 58 CD children after a gluten-free diet. AE was positive in all but 1 case of CD patients (sensitivity = 97%); false positive results were found in 1/34 controls (specificity = 97%). When a specificity of 95% was fixed, the sensitivities were 97% for AE, 83% for AG-IgA, and 63% for AG-IgG; the sensitivities of anti-tTG were 90, 84, 84, and 75% when measured with Eurospital, Medipan, Inova, and Arnika kits respectively. The new AtTG seems to be accurate enough to be proposed as a noninvasive diagnostic tool for CD diagnosis; the 4 kits analyzed showed similar diagnostic efficacy. J. Clin. Lab. Anal. 15:112,115, 2001. © 2001 Wiley-Liss, Inc. [source] Hepatitis C virus risk: a hepatitis C virus related syndromeJOURNAL OF INTERNAL MEDICINE, Issue 5 2000C. Mazzaro Abstract. Mazzaro C, Panarello G, Tesio F, Santini G, Crovatto M, Mazzi G, Zorat F, Tulissi P, Pussini E, Baracetti S, Campanacci L, Pozzato G (Pordenone General Hospital, Pordenone; University of Trieste, School of Medicine, Trieste, Italy). Hepatitis C virus risk: a hepatitis C virus-related syndrome. J Intern Med 2000 247: 535,545. Background. The association between mixed cryoglobulinemia (MC) and hepatitis C virus (HCV) infection has been recently described in many reports. Objective. The aim of this study was to evaluate the long-term prognosis of hepatitis C virus-positive patients affected by mixed cryoglobulinemia with or without kidney involvement. Patients. At total of 119 hepatitis C virus-positive patients affected by mixed cryoglobulinemia were divided in two groups. Group A: mixed cryoglobulinemia without kidney involvement (103 cases); group B: mixed cryoglobulinemia with glomerulonephritis (GN) (16 cases). A further 37 patients affected by mesangio-proliferative glomerulonephritis (MPGN) were evaluated as controls (group C). Methods. Anti-hepatitis C virus antibodies were determined by commercial kits and hepatitis C virus-RNA was detected by polymerase chain reaction (PCR) amplification of the 5, untranslated region (5,UTR) of the virus. The hepatitis C virus genotype was determined according to Okamoto. Liver biopsy was performed in 62 patients, bone marrow biopsy in 65 patients, and kidney biopsy in all patients with proteinuria. Results. In group A, 46 patients (45%) were affected by chronic liver disease (CLD), 21 (20%) by low-grade non-Hodgkin's lymphoma (NHL) and 16 (15%) by both diseases. All patients of group B were affected by type I membrano-proliferative glomerulonephritis, 3 (19%) by chronic liver disease, 6 (37%) by low-grade non-Hodgkin's lymphoma, and 7 (44%) by both diseases. Several genotypes of hepatitis C virus were found, but Type 1b was prevalent. In group C, no patient showed chronic liver disease or non-Hodgkin's lymphoma. Younger age, higher mean blood pressure, lower C4 serum level, and poorer survival significantly distinguished group B from group A. Survival rates at 5 years were: 87.4% for group A, 89.5% for group C, and 50.0% for group B. None of the patients of group B developed kidney failure requiring dialysis, whilst infections were the leading cause of death. Conclusions. In hepatitis C virus-positive patients, the presence of mixed cryoglobulinemia associated with kidney involvement seems to indicate a new syndrome characterized by immune system impairment, lack of progression to kidney failure, and poor survival (hepatitis C virus-Risk syndrome). [source] HTLV-II infection associated with a chronic neurodegenerative disease: Clinical and molecular analysisJOURNAL OF MEDICAL VIROLOGY, Issue 2 2002Edimilson A. Silva Abstract HTLV II is a retrovirus endemic in some Amerindian tribes and spread worldwide with a high prevalence among intravenous drug abusers. It has three different genetic subtypes a, b, and d, defined mainly by the long terminal repeat (LTR) region. HTLV II has been associated with a neurodegenerative disease in few cases. We describe the first well-documented case in Brazil where the virus is endemic in isolated ethnic groups. The patient is a 55-year-old woman with a chronic and painful syndrome characterized by spastic paraparesis, hyperactive reflexes and spastic bladder. Somatosensory evoked potential indicates a thoracic spinal cord lesion. Computer tomography showed periventricular demyelination. Enzyme-linked immunosorbent assay was positive for HTLV I/II whereas the discriminatory Western blot was indeterminate. Molecular analysis of the Tax region revealed a HTLV II pattern that was also confirmed through sequencing the LTR region. Phylogenetic analysis of the LTR sequence shows an HTLV IIa subtype that clustered with the virus isolated from Kayapo Indians and Brazilian urban intravenous drug users. Indeterminate Western blots are frequently found using commercial kits, therefore we recommend that all cases in which a myelopathy is associated with an indeterminate serological result should be evaluated by PCR to determine the actual number of HTLV II associated myelopathy cases. J. Med. Virol. 66:253,257, 2002. © 2002 Wiley-Liss, Inc. [source] Sensitivity of functional protein S assays to protein S deficiency: a comparative study of three commercial kitsJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 5 2003I. Jennings No abstract is available for this article. [source] Performance of sequence analysis, INNO-LiPA line probe assays and AFFIGENE assays in the detection of hepatitis B virus polymerase and precore/core promoter mutationsJOURNAL OF VIRAL HEPATITIS, Issue 6 2006A. Olivero Summary., In this study, we compare results obtained by sequences analysis and commercial kits in the detection of hepatitis B virus (HBV) polymerase and precore (PC) and core promoter mutations. A total of 23 serum samples from lamivudine treated patients were tested for polymerase mutations by direct sequencing, INNO-LiPA HBV DR and AFFIGENE HBV DE/3TC. Full concordance among the three assays was observed in 63% of the total analysed codons. Concordant results were obtained between sequencing and LiPA in 80%, between sequencing and AFFIGENE in 73% and between LiPA and AFFIGENE in 74% of all tested codons. All discrepancies were observed in mixed population samples in which AFFIGENE and LiPA detected additional viral variants not revealed by sequence. In two patients, with serial samples, LiPA detected earlier than sequence and AFFIGENE an emerging mutate strain. PC and core promoter viral variants were detected in 28 serum samples collected from 14 HBV inactive carriers and from 14 hepatitis B patients with chronic liver disease. Direct sequencing, INNO-LiPA HBV PreCore and AFFIGENE HBV MUTANT VL 19 showed fully coincident results in 88% of tested positions. These findings showed that all assays evaluated were sensitive and accurate tools to analyse HBV genomic variability. Sequence analysis is essential to study new emerging mutations as LiPA and AFFIGENE assays are more easily useful in clinical laboratories to detect the appearance of well-characterized HBV variants. [source] Skin testing for immediate hypersensitivity to betalactams: comparison between two commercial kitsALLERGY, Issue 8 2006J. L. Rodríguez-Bada Introduction:, Skin testing with major and minor determinants of benzylpenicillin is the recommended standard practice to evaluate subjects with immediate hypersensitivity to betalactams. The withdrawal of these products from the market has set us back to the early days, before the introduction of reagents for in vivo testing. Objectives:, To compare a recently released kit of benzylpenicillin conjugated to poly- l -lysine (PPL) and minor determinants mixture (MDM) with the previously existing kit in a positive control group of subjects sensitized to major and/or minor determinants of benzylpenicillin. Methods:, Skin tests with both kits were made in a group of positive subjects previously diagnosed with immediate hypersensitivity to penicillins and with positive results to PPL and/or MDM and in a negative control group. Radioallergosorbent test (RAST) inhibition assays with a pool of sera and individual samples were carried out to compare the inhibition capacity of PPL and MDM of both kits. Results:, Of 22 cases selected from our historical group, 14 were positive: eight to PPL, three to MDM and three to both. These results were equivalent for both kits. RAST inhibition studies showed similar potencies in the inhibition of PPL and MDM. Conclusions:, Both tests show similar results in terms of RAST inhibition assays and skin tests sensitivity and specificity in the groups selected. The new assay can be used for the same purpose and indications as the previous test. [source] An inexpensive, automation-friendly protocol for recovering high-quality DNAMOLECULAR ECOLOGY RESOURCES, Issue 4 2006NATALIA V. IVANOVA Abstract Although commercial kits are available for automated DNA extraction, ,artisanal' protocols are not. In this study, we present a silica-based method that is sensitive, inexpensive and compliant with automation. The effectiveness of this protocol has now been tested on more than 5000 animal specimens with highly positive results. [source] Yeast associated with human infections in south-eastern NigeriaMYCOSES, Issue 6 2006L. N. Abia-Bassey Summary A total of 1921 specimens from nine clinical sources were examined by direct microscopy and culture to recover yeast associated with human infection. Identification of yeast was based on their carbon assimilation patterns, using API 20C AUX and ID 32 C (bioMérieux, France) commercial kits. A total of 178 specimens (9.3%) were positive for yeast. Most of the yeast isolates were recovered from urine samples and genital swabs. Prevalence was significantly higher in women (14.7%) than in men (1.4%) (P < 0.05). The age group 21,30 years recorded the highest prevalence of yeast infection (65.2%) followed by age group 11,20 years (16.9%) and >40 years (9.0%). When genital samples were considered, prevalence was significantly higher in the age group 21,30 years than that in older ones (P < 0.05). Isolates recovered included seven species of Candida and Trichosporon inkin. C. albicans accounted for the highest number of isolates (128) followed by C. tropicalis (23) and C. parapsilosis (9). Two isolates each of C. famata and C. norvegensis were recorded and are reported for the first time in Nigeria. The two isolates of T. inkin were recovered from perianal lesions and are also reported for the first time from Nigeria. C. albicans, C. glabrata, C. parapsilosis and C. krusei were found to be the most common yeast species that act as agents of human disease in south-eastern Nigeria. [source] Effect of vitamin C on oxidative liver injury due to isoniazid in ratsPEDIATRICS INTERNATIONAL, Issue 1 2010Yakup Ergul Abstract Background:, The aim of the present study was to investigate the effect of different doses of vitamin C on oxidative liver injury due to isoniazid (INH) in rats. Methods:, Rats were divided into four subgroups, each containing 10 rats. Group 1 was the control group; group 2, INH 50 mg/kg per day; group 3, INH 50 mg/kg per day + low-dose vitamin C (100 mg/kg per day); group 4, INH 50 mg/kg per day + high-dose vitamin C (1000 mg/kg per day). INH and vitamin C were administered into their stomachs through an oral tube. After 21 days, measurements were made in both serum and homogenized liver tissues. The levels of glutathione (GSH), superoxide dismutase (SOD) and other biochemical variables were measured. Malondialdehyde (MDA), glutathione peroxidase (GSH-px) and vitamin C were measured using commercial kits. Results:, Aspartate amino transferase and alanine aminotransferase in group 2 were higher than those in groups 1, 3 and 4 (P < 0.008 for both). Serum and tissue levels of MDA in group 2 were higher than that in groups 1 and 3 (P < 0.008 for both). There was no difference in the SOD levels between the four groups (P= 0.095). Erythrocyte and tissue GSH in group 2 were higher than that in groups 1 and 3 (P < 0.008 for both). Interestingly, erythrocyte and tissue GSH in group 4 were lower than those in group 1 (P < 0.008 for both). Erythrocyte level of GSH-px in group 2 was higher than that in groups 1 and 3 (P < 0.008 for both). Conclusions:, INH-induced liver injury is associated with oxidative stress, and co-administration of low-dose vitamin C may reduce this damage effectively in a rat model. The antioxidant effect of high-dose vitamin C does not seem more potent compared to the low dose. [source] Production of lipid peroxidation products in osteoarthritic tissues: New evidence linking 4-hydroxynonenal to cartilage degradation,ARTHRITIS & RHEUMATISM, Issue 1 2006Barbara Morquette MSc Objective The lipid peroxidation product 4-hydroxynonenal (HNE) is prominently produced in osteoarthritic (OA) synovial cells, but its specific contribution to cartilage destruction is not understood. This study was designed to test whether HNE signaling and binding are involved in OA cartilage degradation through type II collagen (CII) and matrix metalloproteinase 13 (MMP-13) modulation. Methods HNE levels in synovial fluid and in isolated OA chondrocytes treated with free radical donors were determined by enzyme-linked immunosorbent assay. The formation of the HNE/CII adducts was measured in cartilage explants by immunoprecipitation. Levels of CII and MMP-13 messenger RNA and protein were determined by reverse transcription,polymerase chain reaction, Western blotting, and by the use of commercial kits. Results Levels of HNE/protein adducts were higher in OA synovial fluid compared with normal synovial fluid and were higher in OA chondrocytes treated with free radical donors compared with untreated cells. In cartilage explants, HNE induced CII cleavage, as established by the generation of neoepitopes. The level of HNE/CII adducts was increased in OA cartilage explants incubated with free radical donors. Modification of CII by HNE accelerated its degradation by active MMP-13. In isolated OA chondrocytes, HNE inhibited the expression of CII and tissue inhibitor of metalloproteinases 1 and induced MMP-13 mainly through activation of p38 MAPK. In vitro, HNE binding to MMP-13 activated this enzyme at a molar ratio of 1:100 (MMP-13 to HNE). Conclusion The increased level of HNE in OA cartilage and the ability of HNE to induce transcriptional and posttranslational modifications of CII and MMP-13 suggest that this aldehyde could play a role in OA. [source] Age-specific reference levels of serum prostate-specific antigen and prostate volume in healthy Arab menBJU INTERNATIONAL, Issue 3 2005Elijah O. Kehinde OBJECTIVE To determine age-specific reference ranges for serum prostate-specific antigen (PSA) concentration and prostate volumes in a population of healthy Arab men. SUBJECTS AND METHODS Blood samples were taken from 396 healthy Arab men (from Kuwait and Oman) aged 15,79 years and from across the social spectrum. Men aged >40 years had a digital rectal examination and transrectal ultrasonography of the prostate to determine prostate volume. The serum PSA level was measured using commercial kits, and age-specific ranges for PSA levels and prostate volume determined. RESULTS The serum PSA ranges (ng/mL) for each age range in Arab men were: 40,49 years, 0,0.9; 60,69, 0,2.7; 70,79, 0,5.5 ng/mL; the respective prostate volumes were 8,22, 9,30 and 10,33 mL. The serum PSA level and prostate volume correlated with age (P < 0.001). Arab men had lower serum PSA levels and prostate volumes than those reported for Caucasians, but similar to those reported for Asians (Japanese and Chinese). CONCLUSION These results indicate that Arab men have lower PSA levels and prostate volumes than Caucasians. The levels are slightly lower than those reported in the Japanese and, as in the Japanese, low PSA levels and small prostate volumes might be related to the low incidence of clinical prostate cancer in Arab men. [source] A method for fast and simple detection of major diarrhoeagenic Escherichia coli in the routine diagnostic laboratoryCLINICAL MICROBIOLOGY AND INFECTION, Issue 5 2007S. Persson Abstract A multiplex PCR was developed for the detection of the following genes characteristic of diarrhoeagenic Escherichia coli (DEC): verocytotoxins 1 (vtx1) and 2 (vtx2), characteristic of verocytotoxin-producing E. coli (VTEC); intimin (eae), found in enteropathogenic E. coli (EPEC), attaching and effacing E. coli and VTEC; heat-stable enterotoxin (estA) and heat-labile enterotoxin (eltA), characteristic of enterotoxigenic E. coli (ETEC); and invasive plasmid antigen (ipaH), characteristic of enteroinvasive E. coli (EIEC) and Shigella spp. The method allowed the simultaneous identification of all six genes in one reaction, and included a 16S rDNA internal PCR control. When applied to pure cultures from a reference strain collection, all virulence genes in 124 different DEC strains and 15 Shigella spp. were identified correctly, and there were no cross-reactions with 13 non- E. coli species. The detection limit of the method was 102,103 DEC CFU/PCR in the presence of 106 non-target cells. When the multiplex PCR was tested with colonies from plate cultures of clinical stool samples, it was a faster, more sensitive, less expensive and less laborious diagnostic procedure than DNA hybridisation. When used with DNA purified from spiked stool samples (by two different commercial kits), the method had a detection limit of 106 CFU/mL stool sample. [source] Comparison of the performance of serological kits for Helicobacter pylori infection with European and Asian study populationsCLINICAL MICROBIOLOGY AND INFECTION, Issue 11 2006T. T. H. Hoang Abstract Most commercial kits for the detection of Helicobacter pylori were developed and validated with Western populations, and some have been found to perform less well with Asian populations. This study compared the performances of three serological kits with Swedish and Vietnamese peptic ulcer patients and asymptomatic individuals. The Pyloriset EIA-GIII and HM-CAP ELISA kits indicated that Asian populations had lower antibody titres to H. pylori than European populations. Despite the difference, the Pyloriset EIA-GIII kit performed well with Vietnamese peptic ulcer patients and population controls. The HM-CAP ELISA kit had a significantly lower performance with Asian populations that could not be improved by adjustments to the cut-off level. The Helicoblot 2.1 immunoblot kit performed equally well with Vietnamese and Swedish populations, although the response rate to the 35-kDa band was significantly lower with Vietnamese individuals. [source] | |||||||||||||