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Sample records for interferon-gamma release assays

  1. An interferon-gamma release assay test performs well in routine screening for tuberculosis

    DEFF Research Database (Denmark)

    Vestergaard Danielsen, Allan; Fløe, Andreas; Lillebæk, Troels

    2014-01-01

    Introduction: A positive interferon-gamma release assay (IGRA) is regarded as proof of latent Mycobacterium tuberculosis infection. We conducted an evaluation of the IGRA test “T-SPOT.TB” to test its performance during clinical routine use by analysing the positivity rate and odds, effect of season...... and sensitivity. Material and methods: Data from T-SPOT.TB testing together with age and test indications (anti-tumour necrosis factor alpha (TNFα) candidate, contact investigation or suspicion of tuberculosis (TB)) were combined with mycobac­teria culture results. Results: A total of 1,809 patients were tested....... Conclusive results were achieved for 1,780 patients (98.4%). Among these, 4.6% of anti-TNFα candidates, 19.3% of contacts and 24.4% of TB suspects tested positive. Compared with anti-TNFα candidates, the odds for a positive result were significantly higher for contact investigations (odds ratio (OR), mean...

  2. Executive Summary of the Guidelines for the Use of interferon-gamma Release Assays in the Diagnosis of Tuberculosis Infection.

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    Santin, Miguel; García-García, José-María; Rigau, David; Altet, Neus; Anibarro, Luis; Casas, Irma; Díez, Nuria; García-Gasalla, Mercedes; Martínez-Lacasa, Xavier; Penas, Antón; Pérez-Escolano, Elvira; Sánchez, Francisca; Domínguez, José

    2016-09-01

    Interferon-gamma release assays are widely used for the diagnosis of tuberculosis infection in Spain. However, there is no consensus on their application in specific clinical scenarios. To develop a guide-line for their use, a panel of experts comprising specialists in infectious diseases, respiratory diseases, microbiology, pediatrics and preventive medicine, together with a methodologist, conducted a systematic literature search, summarized the findings, rated the quality of the evidence, and formulated recommendations following the Grading of Recommendations of Assessment Development and Evaluations methodology. This document provides evidence-based guidance on the use of interferon-gamma release assays for the diagnosis of tuberculosis infection in patients at risk of tuberculosis or suspected of having active disease. The guidelines will be applicable to specialist and primary care, and public health. Copyright © 2016 SEPAR. Publicado por Elsevier España, S.L.U. All rights reserved.

  3. Frequency of indeterminate results from an interferon-gamma release assay among HIV-infected individuals

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    Sandra Maria do Valle Leone de Oliveira

    Full Text Available ABSTRACT Objective: To evaluate the frequency of and factors associated with indeterminate interferon-gamma release assay (IGRA results in people living with HIV/AIDS (PLWHA. Methods: We tested 81 PLWHA in the central-west region of Brazil, using the tuberculin skin test and an IGRA. Information on sociodemographic and clinical variables was gathered through the use of questionnaires and from medical records. The association of those variables with indeterminate results was analyzed by calculating the adjusted ORs in a multivariate logistic regression model. Concordance was evaluated by determining the kappa statistic. Results: Among the 81 patients evaluated, the tuberculin skin test results were positive in 18 (22.2% of the patients, and the IGRA results were positive in 10 (12.3%, with a kappa of 0.62. The IGRA results were indeterminate in 22 (27.1% of the patients (95% CI: 17.8-38.1%. The odds of obtaining indeterminate results were significantly higher in smokers (adjusted OR = 6.0; 95% CI: 1.4-26.7 and in samples stored for less than 35 days (adjusted OR = 14.0; 95% CI: 3.1-64.2. Patients with advanced immunosuppression (CD4+ T-cell count < 200 cells/mm3 were at a higher risk for indeterminate results (OR adjusted for smoking and inadequate duration of sample storage = 4.7; 95% CI: 0.91-24.0, although the difference was not significant. Conclusions: The high prevalence of indeterminate results can be a major limitation for the routine use of IGRAs in PLWHA. The need to repeat the test increases its costs and should be taken into account in cost-effectiveness studies. The processing of samples can significantly alter the results.

  4. Reproducibility of Interferon Gamma (IFN-γ) Release Assays. A Systematic Review

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    Tagmouti, Saloua; Slater, Madeline; Benedetti, Andrea; Kik, Sandra V.; Banaei, Niaz; Cattamanchi, Adithya; Metcalfe, John; Dowdy, David; van Zyl Smit, Richard; Dendukuri, Nandini

    2014-01-01

    Rationale: Interferon gamma (IFN-γ) release assays for latent tuberculosis infection result in a larger-than-expected number of conversions and reversions in occupational screening programs, and reproducibility of test results is a concern. Objectives: Knowledge of the relative contribution and extent of the individual sources of variability (immunological, preanalytical, or analytical) could help optimize testing protocols. Methods: We performed a systematic review of studies published by October 2013 on all potential sources of variability of commercial IFN-γ release assays (QuantiFERON-TB Gold In-Tube and T-SPOT.TB). The included studies assessed test variability under identical conditions and under different conditions (the latter both overall and stratified by individual sources of variability). Linear mixed effects models were used to estimate within-subject SD. Measurements and Main Results: We identified a total of 26 articles, including 7 studies analyzing variability under the same conditions, 10 studies analyzing variability with repeat testing over time under different conditions, and 19 studies reporting individual sources of variability. Most data were on QuantiFERON (only three studies on T-SPOT.TB). A considerable number of conversions and reversions were seen around the manufacturer-recommended cut-point. The estimated range of variability of IFN-γ response in QuantiFERON under identical conditions was ±0.47 IU/ml (coefficient of variation, 13%) and ±0.26 IU/ml (30%) for individuals with an initial IFN-γ response in the borderline range (0.25–0.80 IU/ml). The estimated range of variability in noncontrolled settings was substantially larger (±1.4 IU/ml; 60%). Blood volume inoculated into QuantiFERON tubes and preanalytic delay were identified as key sources of variability. Conclusions: This systematic review shows substantial variability with repeat IFN-γ release assays testing even under identical conditions, suggesting that reversions

  5. The role of interferon gamma release assays in the monitoring of response to anti-tuberculosis treatment in children.

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    Shaik, Junaid; Pillay, Manormoney; Jeena, Prakash

    2014-09-01

    Successful control of childhood TB requires early diagnosis, effective chemotherapy and a method of evaluating the response to therapy. Identification of suitable biomarkers that predict the response to anti-TB therapy may allow the duration of treatment to be shortened. The majority of biomarker studies in paediatric TB have focused on the role of T cell-based interferon-gamma (IFN-γ) release assays (IGRAs) in the diagnosis of either latent or active disease. Little has been published on the role of IGRAs in the monitoring response to therapy in children. We reviewed the available literature to ascertain the value of IGRAs in the monitoring of response to anti-TB therapy in children. We explored the results of the few studies that have investigated the role of IGRAs as markers of response to anti-TB treatment in children. We conclude that the role of IGRAs as surrogate markers appears promising. Robust clinical trials are, however, needed to entrench the value of IGRAs as surrogate biomarkers of response to anti-TB therapy in children. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. Private sector tuberculosis prevention in the US: Characteristics associated with interferon-gamma release assay or tuberculin skin testing.

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    Stockbridge, Erica L; Miller, Thaddeus L; Carlson, Erin K; Ho, Christine

    2018-01-01

    To determine whether latent tuberculosis infection risk factors are associated with an increased likelihood of latent tuberculosis infection testing in the US private healthcare sector. A national sample of medical and pharmacy claims representing services rendered January 2011 through December 2013 for 3,997,986 commercially insured individuals in the US who were 0 to 64 years of age. We used multivariable logistic regression models to determine whether TB/LTBI risk factors were associated with an increased likelihood of Interferon-Gamma Release Assay (IGRA) or Tuberculin Skin Test (TST) testing in the private sector. 4.31% (4.27-4.34%) received at least one TST/IGRA test between 2011 and 2013 while 1.69% (1.67-1.72%) received a TST/IGRA test in 2013. Clinical risk factors associated with a significantly increased likelihood of testing included HIV, immunosuppressive therapy, exposure to tuberculosis, a history of tuberculosis, diabetes, tobacco use, end stage renal disease, and alcohol use disorder. Other significant variables included gender, age, asthma, the state tuberculosis rate, population density, and percent of foreign-born persons in a county. Private sector TST/IGRA testing is not uncommon and testing varies with clinical risk indicators. Thus, the private sector can be a powerful resource in the fight against tuberculosis. Analyses of administrative data can inform how best to leverage private sector healthcare toward tuberculosis prevention activities.

  7. Serial interferon-gamma release assays during treatment of active tuberculosis in young adults

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    Lee Choon-Taek

    2010-10-01

    Full Text Available Abstract Background The role of interferon-γ release assay (IGRA in monitoring responses to anti-tuberculosis (TB treatment is not clear. We evaluated the results of the QuantiFERON-TB Gold In-tube (QFT-GIT assay over time during the anti-TB treatment of adults with no underlying disease. Methods We enrolled soldiers who were newly diagnosed with active TB and admitted to the central referral military hospital in South Korea between May 1, 2008 and September 30, 2009. For each participant, we preformed QFT-GIT assay before treatment (baseline and at 1, 3, and 6 months after initiating anti-TB medication. Results Of 67 eligible patients, 59 (88.1% completed the study protocol. All participants were males who were human immunodeficiency virus (HIV-negative and had no chronic diseases. Their median age was 21 years (range, 20-48. Initially, 57 (96.6% patients had positive QFT-GIT results, and 53 (89.8%, 42 (71.2%, and 39 (66.1% had positive QFT-GIT results at 1, 3, and 6 months, respectively. The IFN-γ level at baseline was 5.31 ± 5.34 IU/ml, and the levels at 1, 3, and 6 months were 3.95 ± 4.30, 1.82 ± 2.14, and 1.50 ± 2.12 IU/ml, respectively. All patients had clinical and radiologic improvements after treatment and were cured. A lower IFN-γ level, C-reactive protein ≥ 3 mg/dl, and the presence of fever (≥ 38.3°C at diagnosis were associated with negative reversion of the QFT-GIT assay. Conclusion Although the IFN-γ level measured by QFT-GIT assay decreased after successful anti-TB treatment in most participants, less than half of them exhibited QFT-GIT reversion. Thus, the reversion to negativity of the QFT-GIT assay may not be a good surrogate for treatment response in otherwise healthy young patients with TB.

  8. Interferon-gamma release assay and Rifampicin therapy for household contacts of tuberculosis.

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    Wang, Jann-Yuan; Shu, Chin-Chung; Lee, Chih-Hsin; Yu, Chong-Jen; Lee, Li-Na; Yang, Pan-Chyr

    2012-03-01

    Longitudinal studies in household contacts to identify subgroups at risk of active tuberculosis are lacking. Household contacts of pulmonary tuberculosis patients were prospectively enrolled to receive chest radiography, sputum studies, and T-SPOT.TB assay at initial visit. Repeat examinations every 6 months for 3 years, and 4-month rifampin preventive therapy for T-SPOT.TB-positive contacts were provided. We investigated factors predicting T-SPOT.TB-positivity and active pulmonary tuberculosis. 583 contacts were enrolled with a follow-up duration of 20.7 ± 9.4 months. 176 (30.2%) were T-SPOT.TB-positive initially and 32 (18.2%) of them received preventive therapy. Old age, living in the same room/house with the index case, the index case having a high smear grade (3+ ∼ 4+) and pulmonary cavitation were associated with T-SPOT.TB-positivity. Active tuberculosis developed in 9 T-SPOT.TB-positive contacts; risk factors included T-SPOT.TB-positivity without preventive therapy, living in the same room, and the index case being ≤50 years or female. 108 (61.4%) T-SPOT.TB-positive contacts had repeat examinations. Forty-five had T-SPOT.TB reversion and none of them developed active tuberculosis. Household contacts who are T-SPOT.TB-positive and live in the same room as the index case are at risk of active tuberculosis and require preventive therapy and close follow-up. Copyright © 2011 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

  9. Interferon gamma release assays for the diagnosis of latent TB infection in HIV-infected individuals in a low TB burden country.

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    Cheallaigh, Clíona Ní

    2013-01-01

    Interferon gamma release assays (IGRAs) are used to diagnose latent tuberculosis infection. Two IGRAs are commercially available: the Quantiferon TB Gold In Tube (QFT-IT) and the T-SPOT.TB. There is debate as to which test to use in HIV+ individuals. Previous publications from high TB burden countries have raised concerns that the sensitivity of the QFT-IT assay, but not the T-SPOT.TB, may be impaired in HIV+ individuals with low CD4+ T-cell counts. We sought to compare the tests in a low TB burden setting.

  10. The impact of HIV infection and CD4 cell count on the performance of an interferon gamma release assay in patients with pulmonary tuberculosis

    DEFF Research Database (Denmark)

    Aabye, Martine G.; Ravn, Pernille; PrayGod, George

    2009-01-01

    pulmonary tuberculosis (PTB) in a TB- and HIV-endemic population and the effect of HIV-infection and CD4 cell count on test performance. METHODOLOGY/PRINCIPAL FINDINGS: 161 patients with sputum culture confirmed PTB were subjected to HIV- and QFT-IT testing and measurement of CD4 cell count. The QFT......BACKGROUND: The performance of the tuberculosis specific Interferon Gamma Release Assays (IGRAs) has not been sufficiently documented in tuberculosis- and HIV-endemic settings. This study evaluated the sensitivity of the QuantiFERON TB-Gold In-Tube (QFT-IT) in patients with culture confirmed...

  11. Interferon-gamma release assay for the diagnosis of latent tuberculosis infection: A latent-class analysis.

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    Tan N Doan

    Full Text Available Accurate diagnosis and subsequent treatment of latent tuberculosis infection (LTBI is essential for TB elimination. However, the absence of a gold standard test for diagnosing LTBI makes assessment of the true prevalence of LTBI and the accuracy of diagnostic tests challenging. Bayesian latent class models can be used to make inferences about disease prevalence and the sensitivity and specificity of diagnostic tests using data on the concordance between tests. We performed the largest meta-analysis to date aiming to evaluate the performance of tuberculin skin test (TST and interferon-gamma release assays (IGRAs for LTBI diagnosis in various patient populations using Bayesian latent class modelling.Systematic search of PubMeb, Embase and African Index Medicus was conducted without date and language restrictions on September 11, 2017 to identify studies that compared the performance of TST and IGRAs for LTBI diagnosis. Two IGRA methods were considered: QuantiFERON-TB Gold In Tube (QFT-GIT and T-SPOT.TB. Studies were included if they reported 2x2 agreement data between TST and QFT-GIT or T-SPOT.TB. A Bayesian latent class model was developed to estimate the sensitivity and specificity of TST and IGRAs in various populations, including immune-competent adults, immune-compromised adults and children. A TST cut-off value of 10 mm was used for immune-competent subjects and 5 mm for immune-compromised individuals.A total of 157 studies were included in the analysis. In immune-competent adults, the sensitivity of TST and QFT-GIT were estimated to be 84% (95% credible interval [CrI] 82-85% and 52% (50-53%, respectively. The specificity of QFT-GIT was 97% (96-97% in non-BCG-vaccinated and 93% (92-94% in BCG-vaccinated immune-competent adults. The estimated figures for TST were 100% (99-100% and 79% (76-82%, respectively. T-SPOT.TB has comparable specificity (97% for both tests and better sensitivity (68% versus 52% than QFT-GIT in immune-competent adults

  12. Use of interferon-gamma release assays in a health care worker screening program: experience from a tertiary care centre in the United States.

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    Joshi, Manish; Monson, Thomas P; Woods, Gail L

    2012-01-01

    Interferon-gamma release assays including the QuantiFERON-TB Gold In-Tube test (QFT-GIT [Cellestis Ltd, Australia]) may be used in place of the tuberculin skin test (TST) in surveillance programs for Mycobacterium tuberculosis infection control. However, data on performance and practicality of the QFT-GIT in such programs for health care workers (HCWs) are limited. To assess the performance, practicality and reversion rate of the QFT-GIT among HCWs at a tertiary health care institution in the United States. Retrospective chart review of HCWs at Central Arkansas Veterans Healthcare System (Arkansas, USA) who underwent QFT-GIT testing as a part of their employee screening between November 1, 2008 and October 31, 2009. QFT-GIT was used to screen 3290 HCWs. The initial QFT-GIT was interpreted as positive for 129 (3.9%) HCWs, negative for 3155 (95.9%) and indeterminate for six (0.2%). Testing with QFT-GIT was repeated in 45 HCWs who had positive results on the initial test. The QFT-GIT reverted to negative in 18 (40.0%) HCWs, all of whom had negative TST status and initial interferon-gamma values of 0.35 IU⁄mL to 2.0 IU⁄mL. The QFT-GIT test is feasible in large health care setting as an alternative to TST for M tuberculosis infection screening in HCWs but is not free from challenges. The major concerns are the high number of positive test results and high reversion rates on repeat testing, illustrating poor short-term reproducibility of positive QFT-GIT test results. These results suggest adopting a borderline zone between interferon-gamma values of 0.35 IU⁄mL to 2.0 IU⁄mL, and cautious clinical interpretation of values in this range.

  13. Tuberculosis screening of new hospital employees: compliance, clearance to work time, and cost using tuberculin skin test and interferon-gamma release assays.

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    Foster-Chang, Sarah A; Manning, Mary L; Chandler, Laura

    2014-11-01

    Selection of the most suitable test(s) for detection of Mycobacterium tuberculosis (TB) infection should be based on purpose, setting, effectiveness, and cost. Two tests are available to screen for latent TB: the tuberculin skin test (TST) and the more recent interferon-gamma release assays (IGRAs). Based on the administrative, logistic, and technical ease of use, an IGRA trial was initiated by the occupational health department at an urban Veteran's Administration health care facility for TB screening of new employees. As a result, new employees completing the pre-placement process within the organization's designated 14 days increased from 77% to 97%, new employee clearance to work time decreased from 13.18 to 5.91 days, and new employee TB screening costs were reduced by 40%. The IGRA is an acceptable alternative to the TST and has significant potential to improve the process of pre-placement TB screening. Copyright 2014, SLACK Incorporated.

  14. Effect of Pregnancy on Interferon Gamma Release Assay and Tuberculin Skin Test Detection of Latent TB Infection Among HIV-Infected Women in a High Burden Setting.

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    LaCourse, Sylvia M; Cranmer, Lisa M; Matemo, Daniel; Kinuthia, John; Richardson, Barbra A; Horne, David J; John-Stewart, Grace

    2017-05-01

    Peripartum immunologic changes may affect latent tuberculosis infection (LTBI) diagnostic performance among HIV-infected women. HIV-infected women were serially tested with tuberculin skin test (TST) and interferon gamma release assay [QuantiFERON TB Gold In-tube (QFT)] in pregnancy and 6 weeks postpartum in Kenya. Prevalence, sensitivity and agreement, and correlates of QFT/TST positivity were assessed. Quantitative QFT mitogen and Mycobacterium tuberculosis antigen (Mtb-Ag) responses were compared by peripartum stage. Incidence of test conversion at 6 weeks postpartum was evaluated in baseline TST-/QFT- women. Among 100 HIV-infected women, median age was 26 years, median CD4 was 555 cells per cubic millimeter, and 88% were on antiretrovirals. More women were QFT+ than TST+ in both pregnancy (35.4% vs. 13.5%, P = 0.001) and postpartum (29.6% vs. 14.8%, P pregnancy vs. postpartum, and specifically among persistently QFT+ women (Mtb-Ag: 3.46 vs. 4.48 IU/mL, P = 0.007). QFT indeterminate rate was higher in pregnancy (16%) compared with postpartum (0%) because of lower mitogen response. QFT identified >2-fold more women with LTBI compared with TST in pregnancy and postpartum. Lower QFT Mtb-Ag and mitogen responses in pregnancy compared with postpartum suggest that pregnancy-associated immunologic changes may influence LTBI test performance.

  15. Screening of latent tuberculosis infection among health care workers working in Hajj pilgrimage area in Saudi Arabia, using interferon gamma release assay and tuberculin skin test.

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    Bukhary, Zakeya A; Amer, Soliman M; Emara, Magdy M; Abdalla, Mohammad E; Ali, Sahar A

    2018-01-01

    Interferon gamma release assays (IGRA) is highly specific for Mycobacterium tuberculosis and is the preferred test in BCG-vaccinated individuals. The few studies that have screened health care workers (HCWs) in Saudi Arabia for latent tuberculosis infection (LTBI) using IGRA have varied in agreement with the traditional tuberculin skin test (TST). Assess the prevalence of LTBI among HCWs working in the Hajj pilgrimage using IGRA and TST and measuring their agreement. Cross-sectional prospective. Multiple non-tertiary care hospitals. HCWs who worked during the Hajj pilgrimage in Saudi Arabia in December 2015. Data was collected by standarized questionnaire. Samples were drawn and analyzed by standard methods. The prevalence of LTBI among HCW and the agreement by kappa statistic between QFT-GIT and TST. 520 subjects. Nurses accounted for 30.7% of the sample and physicians, 19.2%. The majority were BCG vaccinated (98.5%). There were a total of 56 positive by QFT-GIT and the LTBI rate was 10.8%. In 50 QFT positive/476 TST negative the LTBI rate was 10.5% in discordant tests, and in 6 QFT positive/44 TST positive it was 13.6% in concordant tests. The overall agreement between both tests was poor-83% and kappa was 0.02. LTBI prevalence was associated with longer employment (13.1 [9.2] years). The QFT-GIT positive test was significantly higher in physicians (P=.02) and in HCWs working in chest hospitals 16/76 (21.05%) (P=.001). Agreement between the tests was poor. QFT-GIT detected LTBI when TST was negative in HCWs who had a history of close contact with TB patients. A second step TST was not feasible within 2-3 weeks. None.

  16. Comparison of interferon {gamma} release assays and conventional screening tests before tumour necrosis factor {alpha} blockade in patients with inflammatory arthritis.

    LENUS (Irish Health Repository)

    Martin, J

    2012-02-01

    OBJECTIVE: To compare the performance of two interferon gamma release assays (IGRAs) and conventional screening tests in patients with inflammatory arthritis undergoing screening for latent tuberculosis infection (LTBI) before treatment with anti-tumour necrosis factor alpha (anti-TNFalpha) compounds. METHODS: Successive patients were subjected to conventional LTBI screening, including a tuberculin skin test (TST). The T-SPOT.TB test was performed on all patients and the QuantiFERON-TB Gold test was performed on a large subset. The results of the IGRAs were compared with the results of conventional screening tests. RESULTS: A total 150 patients were evaluated. The majority (57.9%) had rheumatoid arthritis. Previous vaccination with Bacille Calmette-Guerin was confirmed in 82% of patients. No patient had received prior anti-TB treatment. A total of 57 patients (38.0%) had at least one positive conventional risk factor. In contrast, an unequivocally positive T-SPOT.TB test was seen in only 14\\/143 (9.8%). There was 98.2% agreement between the two IGRAs. Statistically significant associations were found between each of the IGRAs and both TST and risk history, but not chest x-ray (CXR). A positive IGRA result was significantly associated with increased age. TB was not reactivated in any patient during the follow-up period. Interpretation: This study suggests that IGRAs may be useful when screening for LTBI before anti-TNFalpha therapy in patients with immune-mediated inflammatory diseases. The observations reported here also highlight the inadequate performance of CXR as a marker of LTBI.

  17. Tuberculin skin testing and treatment modulates interferon-gamma release assay results for latent tuberculosis in migrants.

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    Matthew K O'Shea

    Full Text Available Identifying latent tuberculosis infection (LTBI in people migrating from TB endemic regions to low incidence countries is an important control measure. However, no prospective longitudinal comparisons between diagnostic tests used in such migrant populations are available.To compare commercial interferon (IFN-gamma release assays (IGRAs and the tuberculin skin test (TST for diagnosing LTBI in a migrant population, and the influence of antecedent TST and LTBI treatment on IGRA performance.This cohort study, performed from February to September 2012, assessed longitudinal IGRA and TST responses in Nepalese military recruits recently arrived in the UK. Concomitant T-SPOT.TB, QFT-GIT and TST were performed on day 0, with IGRAs repeated 7 and 200 days later, following treatment for LTBI if necessary.166 Nepalese recruits were prospectively assessed. At entry, 21 individuals were positive by T-SPOT.TB and 8 individuals by QFT-GIT. There was substantial agreement between TST and T-SPOT.TB positives at baseline (71.4% agreement; κ = 0.62; 95% CI:0.44-0.79, but only moderate concordance between positive IGRAs (38.1% agreement; κ = 0.46; 95% CI:0.25-0.67. When reassessed 7 days following TST, numbers of IGRA-positive individuals changed from 8 to 23 for QFT-GIT (p = 0.0074 and from 21 to 23 for T-SPOT.TB (p = 0.87. This resulted in an increase in IGRA concordance to substantial (64.3% agreement; κ = 0.73; 95% CI:0.58-0.88. Thus, in total on day 0 and day 7 after testing, 29 out of 166 participants (17.5% provided a positive IGRA and of these 13 were TST negative. Two hundred days after the study commenced and three months after treatment for LTBI was completed by those who were given chemoprophylaxis, 23 and 21 participants were positive by T-SPOT.TB or QFT-GIT respectively. When individual responses were examined longitudinally within this population 35% of the day 7 QFT-GIT-positive, and 19% T-SPOT.TB-positive individuals, were

  18. Use of an Interferon Gamma Release Assay (IGRA) to test T-cell responsiveness to soluble Leishmania infantum antigen in whole blood of dogs from endemic areas.

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    Zribi, Lilia; El-Goulli, Amel F; Ben-Abid, Meriem; Gharbi, Mohamed; Ben-Sghaier, Ines; Boufaden, Imed; Aoun, Karim; Bouratbine, Aïda

    2017-11-15

    Interferon-Gamma (IFN-γ) Release Assays (IGRAs) are easy tests that allow rapid screening of primed memory T-cells immunity in response to antigen. The aim of this study was to use IGRA to assess IFN-γ release in response to Soluble Leishmania infantum antigen (SLA) in whole blood of dogs living in endemic area of visceral leishmaniasis and to interpret IGRA results according to clinical examination, specific anti-Leishmania humoral response and presence of L. infantum DNA in blood. The study was carried out on 56 dogs living in greater Tunis area. Physical examination, quantitative serology and PCR on blood were used to characterize dogs' status in relation to Leishmania infection and disease. IGRA consisted on testing by ELISA for IFN-γ-secretion in whole blood after a 20-h challenge with SLA. PBS and Phytohemagglutinin (PHA) stimulations were used as controls. Four groups of dogs were characterized: 31 were negative by both serology and PCR, two had doubtful serology, 10 presented no to mild clinical signs but low antibodies levels and 13 were affected by Canine Leishmaniasis (CanL). In seronegative dogs, IGRA was little contributory in 4 puppies (age dogs (median age=72months, IQR: 45-84 months) that didn't respond to PHA stimulation, IGRA was negative in 19 and positive in three animals with lymph node enlargement. In dogs with doubtful serology, IGRA was positive in one dog and negative in the other. In infected dogs with no to mild clinical signs, one dog exhibited high level of IFN-γ in absence of antigenic stimulation and all the other were positive by IGRA. CanL dogs showed variable IGRA results. Negative IGRAs (n=4) were shown in animals with the highest parasitic burden whereas positive IGRAs (n=5) were shown in dogs with negative PCR or low parasitic load. The 4 remaining dogs either didn't respond to PHA (n=2) or showed non-specific secretion in PBS tube (n=2). The results of this study showed that IGRA is a useful new tool that can assess

  19. Interferon Gamma Release Assay versus Tuberculin Skin Testing among Healthcare Workers of Highly Diverse Origin in a Moderate Tuberculosis Burden Country.

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    Sahal Al Hajoj

    Full Text Available Health care workers (HCW's are always at an increased risk of contracting tuberculosis (TB infection. In Saudi Arabia, Interferon Gamma Release Assay (IGRA has not been evaluated as a screening tool for latent TB infection (LTBI among HCW's considering their high demographic diversity. During February 2012 to January 2015 a cross sectional study has been conducted in a tertiary care center with maximum demographically diverse staff population in the capital city-Riyadh. After a short interview and consenting, all the candidates were subjected to tuberculin skin test (TST and QuantiFERON TB gold In-tube test (QFT. A logistic regression analysis was carried out for establishing the associations between putative risk factors and the diagnostic tests. The candidates were classified according to geographical origin and a detailed analysis was conducted on the impact of their origin towards the results of TST and QFT. Of the 1595 candidates enrolled, 90.6% were BCG vaccinated, female (67.9% and mainly nurses (53.2%. Candidates with high risk of suspected or confirmed TB patient exposure were 56.1% and 76.5% of them had <10 year's work experience. TST positivity was observed in 503 (31.5% candidates, while QFT was positive among 399 (25%. Majority of the candidates were non-Saudi (83% and predominantly (52.4% from Western Pacific region. Concordant results were obtained in 14.2% of positive cases and 57.7% negative cases. The disagreements between the two tests were relatively high (kappa co-efficient-0.312±0.026, p value- <0.00001 as TST positive/QFT negative discordance was 54.8% while TST negative/QFT positive discordance was 15.7%. Age of the candidates, BCG vaccination, and South East Asian origin were associated with TST positivity while Occupational TB exposure and geographical origin of the candidates were associated with QFT positivity. A regular follow up on recently TST converted candidates showed no progression to active TB. The putative

  20. Development of a lion-specific interferon-gamma assay.

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    Maas, M; van Kooten, P J S; Schreuder, J; Morar, D; Tijhaar, E; Michel, A L; Rutten, V P M G

    2012-10-15

    The ongoing spread of bovine tuberculosis (BTB) in African free-ranging lion populations, for example in the Kruger National Park, raises the need for diagnostic assays for BTB in lions. These, in addition, would be highly relevant for zoological gardens worldwide that want to determine the BTB status of their lions, e.g. for translocations. The present study concerns the development of a lion-specific IFN-γ assay, following the production and characterization of monoclonal antibodies specific for lion interferon-gamma (IFN-γ). Recombinant lion IFN-γ (rLIFN-γ) was produced in mammalian cells and used to immunize mice to establish hybridoma cell lines producing monoclonal antibodies. These were used to develop a sensitive, lion IFN-γ-specific capture ELISA, able to detect rLIFN-γ to the level of 160 pg/ml. Recognition of native lion IFN-γ was shown in an initial assessment of supernatants of mitogen stimulated whole blood cultures of 11 known BTB-negative lions. In conclusion, the capture ELISA shows potential as a diagnostic assay for bovine tuberculosis in lions. Preliminary results also indicate the possible use of the test for other (feline) species. Copyright © 2012 Elsevier B.V. All rights reserved.

  1. Use of the johnin PPD interferon-gamma assay in control of bovine paratuberculosis

    DEFF Research Database (Denmark)

    Jungersen, Gregers; Mikkelsen, Heidi; Grell, Susanne N.

    2012-01-01

    Although the interferon-gamma (IFN-γ) assay for measurements of cell-mediated immune (CMI) responses to paratuberculosis PPD (johnin) has been available for close to 20 years, the assay has not yet emerged as the long desired test to identify infected animals at an early time point. Among other...

  2. Evaluation of the performance of two tuberculosis interferon gamma release assays (IGRA-ELISA and T-SPOT.TB) for diagnosing Mycobacterium tuberculosis infection.

    Science.gov (United States)

    Wang, Linchuan; Tian, Xu-Dong; Yu, Yan; Chen, Wei

    2018-04-01

    The IGRA-ELISA and T-SPOT.TB are widely used in China. The aim of the study was to evaluate the performance of the two assays in diagnosis Mycobacterium tuberculosis infection. Of the 3727 patients in the study, 204 underwent testing using both the T-SPOT.TB and IGRA-ELISA, 1794 were tested using the T-SPOT.TB only, and 1729 were tested using the IGRA-ELISA only. The positive rate and consistency of the two assays were analyzed, and their sensitivity and specificity for diagnosing active tuberculosis were compared. There were no significant differences in the positive rate between the T-SPOT.TB test (25.8%) and IGRA-ELISA (28.6%), p = .065. The two assays were highly consistent, with a kappa value of 0.852 (p SPOT.TB test were 82.9% (107/129) and 78.6% (1309/1665), respectively, and those of IGRA-ELISA were 81.7% (94/115) and 75.2% (1214/1614), respectively. There were no significant differences in sensitivity (p > .05), but the specificity of the T-SPOT.TB test was slightly higher than that of IGRA-ELISA (p = .023). Both in terms of diagnosing M. tuberculosis infection and ruling out active tuberculosis, the performance of the IGRA-ELISA-a simple, almost labor-free assay that allows simultaneous processing of a very large number of samples-was well-matched with that of T-SPOT.TB test. However, IGRAs cannot be used as the only test to diagnose active tuberculosis. Copyright © 2018 Elsevier B.V. All rights reserved.

  3. The feasibility of the interferon gamma release assay and predictors of discordance with the tuberculin skin test for the diagnosis of latent tuberculosis infection in a remote Aboriginal community.

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    Gonzalo G Alvarez

    Full Text Available The tuberculin skin test (TST is the standard test used to screen for latent TB infection (LTBI in the northern Canadian territory of Nunavut. Interferon gamma release assays (IGRA are T cell blood-based assays to diagnose LTBI. The Bacillus Calmette-Guerin (BCG vaccine is part of the routine immunization schedule in Nunavut. The objective of this study was to test the feasibility, and predictors of discordance between the Tuberculin Skin Test (TST and the IGRA assay in a medically under-serviced remote arctic Aboriginal population.Both the TST and QuantiFERON-TB Gold (Qiagen group IGRA tests were offered to people in their homes as part of a public health campaign aimed at high TB risk residential areas in Iqaluit, Nunavut, Canada. Feasibility was measured by the capacity of the staff to do the test successfully as measured by the proportion of results obtained.In this population of predominantly young Inuit who were mostly BCG vaccinated, the use of IGRA for the diagnosis of LTBI was feasible. IGRA testing resulted in more available test results reaching patients (95.6% vs 90.9% p = 0.02 but took longer (median 8 days (IGRA vs 2 days (TST, p value < 0.0001. 44/256 participants (17.2% had discordant results. Multivariable regression analysis suggested that discordant results were most likely to have received multiple BCG vaccinations (RR 20.03, 95% CI, 3.94-101.82, followed by BCG given post infancy (RR 8.13, 95% CI, 2.54-26.03 and then to a lesser degree when BCG was given in infancy (RR 6.43, 95% CI, 1.72-24.85.IGRA is feasible in Iqaluit, Nunavut, a remote Arctic community. IGRA testing results in more test results available to patients compared to TST. This test could result in fewer patients requiring latent TB treatment among those previously vaccinated with BCG in a region with limited public health human resources.

  4. Diabetes is associated with lower tuberculosis antigen-specific interferon gamma release in Tanzanian tuberculosis patients and non-tuberculosis controls

    DEFF Research Database (Denmark)

    Faurholt-Jepsen, Daniel; Aabye, Martine Grosos; Jensen, Andreas Vestergaard

    2014-01-01

    in diabetes patients and therefore the validity of interferon gamma release assays (IGRA) may be compromised. The aim of the present study was to assess the association between diabetes and Mycobacterium tuberculosis (Mtb) antigen-specific interferon gamma (IFN-γ) release in a TB endemic area among culture......-confirmed TB patients and non-TB controls. Methods: Culture-confirmed pulmonary TB patients (n = 187) and healthy non-TB neighbourhood controls (n = 190) from Mwanza, Tanzania were tested for the presence of circulating T cells recognizing Mtb antigens using an IGRA. The diabetes status of all participants...

  5. Development of a lion-specific interferon-gamma assay

    NARCIS (Netherlands)

    Maas, M.; Kooten, van P.J.S.; Schreuder, J.; Morar, D.; Tijhaar, E.; Michel, A.L.; Rutten, V.P.M.G.

    2012-01-01

    The ongoing spread of bovine tuberculosis (BTB) in African free-ranging lion populations, for example in the Kruger National Park, raises the need for diagnostic assays for BTB in lions. These, in addition, would be highly relevant for zoological gardens worldwide that want to determine the BTB

  6. Whole blood interferon-gamma assay for baseline tuberculosis screening among Japanese healthcare students.

    Directory of Open Access Journals (Sweden)

    Katsuyuki Hotta

    Full Text Available BACKGROUND: The whole blood interferon-gamma assay (QuantiFERON-TB-2G; QFT has not been fully evaluated as a baseline tuberculosis screening test in Japanese healthcare students commencing clinical contact. The aim of this study was to compare the results from the QFT with those from the tuberculin skin test (TST in a population deemed to be at a low risk for infection with Mycobacterium tuberculosis. METHODOLOGY/PRINCIPAL FINDINGS: Healthcare students recruited at Okayama University received both the TST and the QFT to assess the level of agreement between these two tests. The interleukin-10 levels before and after exposure to M tuberculosis-specific antigens (early-secreted antigenic target 6-kDa protein [ESAT-6] and culture filtrate protein 10 [CFP-10] were also measured. Of the 536 healthcare students, most of whom had been vaccinated with bacillus-Calmette-Guérin (BCG, 207 (56% were enrolled in this study. The agreement between the QFT and the TST results was poor, with positive result rates of 1.4% vs. 27.5%, respectively. A multivariate analysis also revealed that the induration diameter of the TST was not affected by the interferon-gamma concentration after exposure to either of the antigens but was influenced by the number of BCG needle scars (p = 0.046. The whole blood interleukin-10 assay revealed that after antigen exposure, the median increases in interleukin-10 concentration was higher in the subgroup with the small increase in interferon-gamma concentration than in the subgroup with the large increase in interferon-gamma concentration (0.3 vs. 0 pg/mL; p = 0.004. CONCLUSIONS/SIGNIFICANCE: As a baseline screening test for low-risk Japanese healthcare students at their course entry, QFT yielded quite discordant results, compared with the TST, probably because of the low specificity of the TST results in the BCG-vaccinated population. We also found, for the first time, that the change in the interleukin-10 level after exposure to

  7. Concordancia de las pruebas de tuberculina e Interferón gamma en población reclusa Concordance of tuberculin tests and Interferon gamma release assays in the prison population

    Directory of Open Access Journals (Sweden)

    A. Marco Mouriño

    2011-06-01

    Full Text Available Objetivos: Estudiar en población penitenciaria la concordancia de la prueba de la tuberculina (PT y las pruebas de interferón gamma (IFG. Material y métodos: Estudio prospectivo realizado en una prisión en mayo-junio de 2009. Se estudian los ingresos sin antecedente de tuberculosis (TB o con PT previa negativa o no realizada. Se realizó IDR de Mantoux (positivo ³ 10 mm y extracción sanguínea para prueba de IFG (QuantiFERON®-TB Gold. En los infectados, se realizó despistaje de TB. Se pasó un cuestionario y se solicitó consentimiento informado. El estudio fue aprobado por un Comité ético ajeno a instituciones penitenciarias. La concordancia entre PT e IFG se basó en el índice Kappa. Resultados: Se incluyeron 181 casos. El 62% eran extranjeros, el 17% vacunados por BCG, el 8,4% UDI y el 4% VIH+. En los extranjeros había más vacunados, menos UDI y menos infectados por VIH que en autóctonos (p=0,02, p=0,02, y p=0,01, respectivamente. La PT fue positiva en el 24% y la IFG en el 26%. Hubo información de ambas en 149 (82% casos. El 15,8% fueron discordantes. El índice Kappa fue de 0,6 (0,4-0,7. La concordancia varió según subgrupos, siendo mayor en autóctonos (kappa= 0,8 y menor en vacunados (kappa=0,4 e inmigrantes (kappa=0,5. Conclusión: La concordancia global fue moderada-buena, pero en vacunados e inmigrantes fue menor. El nivel de discordancia aconseja ampliar el estudio, así como evaluar que prueba predice mejor el riesgo de progresión a TB y el coste-beneficio de ambas en la población reclusa de nuestro país.Objective: To study the agreement of Tuberculin Skin Tests (TST and Interferon Gamma Release Assays (IGRA when screening tuberculosis infection amongst inmates recently admitted to prison. Materials and methods: Prospective study conducted in a prison during the months of May and June 2009. Inmates without a TB history, with previous TST negatives or without prior TSTs were included. Participants signed an

  8. Use of Novel Recombinant Antigens in the Interferon Gamma Assay for Detection of Mycobacterium Avium Subsp. Paratuberculosis Infection in Cattle

    DEFF Research Database (Denmark)

    Mikkelsen, Heidi; Aagaard, Claus; Nielsen, Søren Saxmose

    2012-01-01

    of the study were to evaluate immunogenicity and specificity of 14 novel recombinant antigens for use in the IFN-γ assay and to assess the consistency of IFN-γ responses. The antigens used were 4 ESAT-6 family members, 4 latency proteins, 4 secreted proteins including Ag85B, 3 other antigens and PPDj......Early stage Mycobacterium avium subsp. paratuberculosis (MAP) infection can be detected by measuring antigen specific cell mediated immune responses by the interferon gamma (IFN-γ) assay. Available IFN-γ assay use purified protein derivate of Johnin (PPDj) leading to low specificity. The objectives...... of the infected and non-infected herds were significantly (Passay using PPDj did not correlate with the results using the novel antigens since 5 of the 17 animals that were positive to PPDj were...

  9. Interferon gamma, interferon-gamma-induced-protein 10, and tuberculin responses of children at high risk of tuberculosis infection

    DEFF Research Database (Denmark)

    Petrucci, Roberta; Abu Amer, Nabil; Gurgel, Ricardo Queiroz

    2008-01-01

    BACKGROUND: Children in contact with adults with pulmonary tuberculosis (TB) are at risk for infection and disease progression, and chemoprophylaxis may reduce this risk. The identification of infection is based on the tuberculin skin test (TST) and interferon-gamma (INF-gamma) release assays. Ot...

  10. Interferon gamma, interferon-gamma-induced-protein 10, and tuberculin responses of children at high risk of tuberculosis infection

    DEFF Research Database (Denmark)

    Petrucci, Roberta; Abu Amer, Nabil; Gurgel, Ricardo Queiroz

    2008-01-01

    BACKGROUND: Children in contact with adults with pulmonary tuberculosis (TB) are at risk for infection and disease progression, and chemoprophylaxis may reduce this risk. The identification of infection is based on the tuberculin skin test (TST) and interferon-gamma (INF-gamma) release assays...

  11. Diagnosis of Coxiella burnetii infection: comparison of a whole blood interferon-gamma production assay and a Coxiella ELISPOT.

    Directory of Open Access Journals (Sweden)

    Teske Schoffelen

    Full Text Available Diagnosis of ongoing or past infection with Coxiella burnetii, the causative agent of Q fever, relies heavily on serology: the measurement of C. burnetii-specific antibodies, reflecting the host's humoral immune response. However, cell-mediated immune responses play an important, probably even more relevant, role in infections caused by the intracellular C. burnetii bacterium. Recent studies have investigated interferon-gamma (IFN-γ based assays, including a whole-blood IFN-γ production assay and a Coxiella enzyme-linked immunospot (Coxiella ELISPOT, as potential diagnostic tools for Q fever diagnosis. Both are in-house developed assays using stimulating antigens of different origin. The main objective of this study was to compare the test performance of the IFN-γ production assay and the Coxiella ELISPOT for detecting a cellular immune response to C. burnetii in Q fever patients, and to assess the correlation between both assays. To that end, both tests were performed in a well-defined patient group of chronic Q fever patients (n = 16 and a group of healthy seronegative individuals (n = 17. Among patients, both the Coxiella ELISPOT and the IFN-γ production assay detected positive response in 14/16. Among controls, none were positive in the Coxiella ELISPOT, whereas the IFN-γ production assay detected positive results in 1/17 and 3/17, when using Henzerling and Nine Mile as stimulating antigens, respectively. These results suggest the Coxiella ELISPOT has a somewhat higher specificity than the IFN-γ production assay when Nine Mile is used as antigen stimulus. The assays showed moderate correlation: the Spearman correlation coefficient r ranged between 0.37-0.60, depending on the antigens used. Further investigation of the diagnostic potential for C. burnetii infection of both assays is warranted.

  12. Interferon Gamma-1b Injection

    Science.gov (United States)

    Interferon gamma-1b injection is used to reduce the frequency and severity of serious infections in people with chronic ... severe, malignant osteopetrosis (an inherited bone disease). Interferon gamma-1b is in a class of medications called ...

  13. Effects of peritoneal fluid from endometriosis patients on interferon-gamma-induced protein-10 (CXCL10) and interleukin-8 (CXCL8) released by neutrophils and CD4+ T cells.

    Science.gov (United States)

    Kim, Ji-Yeon; Lee, Dong-Hyung; Joo, Jong-Kil; Jin, Jun-O; Wang, Ji-Won; Hong, Young-Seoub; Kwak, Jong-Young; Lee, Kyu-Sup

    2009-09-01

    Intraperitoneal immuno-inflammatory changes may be associated with the pathogenesis of endometriosis. We evaluated the effects of peritoneal fluid obtained from patients with endometriosis (ePF) on the release of interferon-gamma-induced protein-10 (IP-10/CXCL10) and interleukin-8 (IL-8/CXCL8) by neutrophils, CD4(+) T cells, and monocytes. Neutrophils, CD4(+) T cells, and monocytes were cultured with ePF and the chemokine levels in the supernatants were then measured using enzyme-linked immunosorbent assay. The addition of ePF to cultures of CD4(+) T cells led to a significant increase in the release of IP-10 when compared with control PF without endometriosis (cPF). There was a positive correlation between the levels of IL-8 and IP-10 in ePF (R = 0.89, P = 0.041), but not between the levels of IP-10 and IL-8 released by neutrophils, CD4(+) T cells, and monocytes. The levels of IP-10 in ePF were positively correlated with the release of IP-10 by ePF-treated neutrophils (R = 0.89, P ePF significantly enhanced the interferon-gamma-induced release of IP-10 by nuetrophils and CD4(+) T cells. These findings suggest that neutrophils and T cells release differential levels of IP-10 and IL-8 in response to stimulation with ePF, and that these cells are a major source of IP-10 in the PF of endometriosis patients.

  14. Development and evaluation of an interferon gamma assay for the diagnosis of tuberculosis in red deer experimentally infected with Mycobacterium bovis.

    Science.gov (United States)

    Risalde, María Ángeles; Thomas, Jobin; Sevilla, Iker; Serrano, Miriam; Ortíz, Jose Antonio; Garrido, Joseba; Domínguez, Mercedes; Domínguez, Lucas; Gortázar, Christian; Ruíz-Fons, Jose Francisco

    2017-11-16

    Red deer (Cervus elaphus) is regarded as an epidemiologically relevant host for Mycobacterium bovis (M. bovis) and closely related members of the Mycobacterium tuberculosis complex that cause animal tuberculosis (TB). The standard antemortem screening test for the detection of TB in deer is the intradermal tuberculin skin test, but the detection of interferon-gamma (IFNγ) produced by white blood cells exposed to M. bovis antigens can be used as an alternative or supplemental assay in most TB eradication/control programs. This study aims to develop an in-house sandwich ELISA for deer IFNγ, based on the cross-reactivity of the antibodies to both cervid and bovine IFNγ, and to evaluate the potential of this assay to detect M. bovis-infected red deer in response to the in vitro stimulation of whole-blood cells with bovine purified protein derivative (bPPD), p22 protein complex derived from bPPD or using the specific tuberculous mycobacterial proteins ESAT-6/CFP-10, Rv3615c and Rv3020c. The positive control stimulant used in this study was pokeweed mitogen, which resulted in a consistent induction of IFNγ in samples from red deer, thus allowing the interpretation of the assay. The percentage of animals correctly classified by this technique as M. bovis non-infected was 100%. The detection of infected animals as positive was high and ranged widely depending upon the antigen and the cut-off value applied, as well as the time after infection. Our findings indicate that this protocol may serve as a reliable assay for the antemortem diagnosis of TB from the initial stage of M. bovis-infection, and may also be adequately sensitive. The suggested optimal antigens and cut-off are bPPD, p22 and the combination of ESAT-6/CFP-10 and Rv3020c with a 0.05 Δ optical density, which yielded a up to 100% correct classification of TB positive and negatve red deer under our experimental conditions. This technique will aid in TB testing of farmed and translocated deer. Future studies

  15. Inhibition of sup 125 I organification and thyroid hormone release by interleukin-1, tumor necrosis factor-alpha, and interferon-gamma in human thyrocytes in suspension culture

    Energy Technology Data Exchange (ETDEWEB)

    Sato, K.; Satoh, T.; Shizume, K.; Ozawa, M.; Han, D.C.; Imamura, H.; Tsushima, T.; Demura, H.; Kanaji, Y.; Ito, Y. (Institute of Clinical Endocrinology, Tokyo (Japan))

    1990-06-01

    To elucidate the mechanism of decreased 131I uptake by the thyroid gland in patients with subacute thyroiditis and painless thyroiditis, human thyroid follicles were cultured with interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF alpha), and/or interferon-gamma (IFN gamma), and the effects of these cytokines on thyroid function were studied in vitro. When human thyrocytes were cultured in RPMI-1640 medium containing 0.5% fetal calf serum and TSH for 5-8 days, the cells incorporated 125I, synthesized de novo (125I)iodotyrosines and (125I)iodothyronines, and secreted (125I)T4 and (125I)T3 into the medium. IL-1 alpha and IL-1 beta inhibited 125I incorporation and (125I)iodothyronine release in a concentration-dependent manner. The minimal inhibitory effect was detected at 10 pg/ml. Electron microscopic examination revealed a marked decrease in lysosome formation in IL-1-treated thyrocytes. TNF alpha and IFN gamma also inhibited thyroid function in a concentration-dependent manner. Furthermore, when thyrocytes were cultured with IL-1, TNF alpha and IFN gamma, these cytokines more than additively inhibited thyroid function. Although the main mechanism of 131I uptake suppression in the thyroid gland in subacute thyroiditis is due to cellular damage and suppression of TSH release, our present findings suggest that IL-1, TNF alpha, and IFN gamma produced in the inflammatory process within the thyroid gland further inhibit iodine incorporation and at least partly account for the decreased 131I uptake by the thyroid gland in destruction-induced hyperthyroidism.

  16. Inhibition of 125I organification and thyroid hormone release by interleukin-1, tumor necrosis factor-alpha, and interferon-gamma in human thyrocytes in suspension culture

    International Nuclear Information System (INIS)

    Sato, K.; Satoh, T.; Shizume, K.; Ozawa, M.; Han, D.C.; Imamura, H.; Tsushima, T.; Demura, H.; Kanaji, Y.; Ito, Y.

    1990-01-01

    To elucidate the mechanism of decreased 131I uptake by the thyroid gland in patients with subacute thyroiditis and painless thyroiditis, human thyroid follicles were cultured with interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF alpha), and/or interferon-gamma (IFN gamma), and the effects of these cytokines on thyroid function were studied in vitro. When human thyrocytes were cultured in RPMI-1640 medium containing 0.5% fetal calf serum and TSH for 5-8 days, the cells incorporated 125I, synthesized de novo [125I]iodotyrosines and [125I]iodothyronines, and secreted [125I]T4 and [125I]T3 into the medium. IL-1 alpha and IL-1 beta inhibited 125I incorporation and [125I]iodothyronine release in a concentration-dependent manner. The minimal inhibitory effect was detected at 10 pg/ml. Electron microscopic examination revealed a marked decrease in lysosome formation in IL-1-treated thyrocytes. TNF alpha and IFN gamma also inhibited thyroid function in a concentration-dependent manner. Furthermore, when thyrocytes were cultured with IL-1, TNF alpha and IFN gamma, these cytokines more than additively inhibited thyroid function. Although the main mechanism of 131I uptake suppression in the thyroid gland in subacute thyroiditis is due to cellular damage and suppression of TSH release, our present findings suggest that IL-1, TNF alpha, and IFN gamma produced in the inflammatory process within the thyroid gland further inhibit iodine incorporation and at least partly account for the decreased 131I uptake by the thyroid gland in destruction-induced hyperthyroidism

  17. Optimization of interferon gamma ELISPOT assay to detect human cytomegalovirus specific T-cell responses in solid organ transplants.

    Science.gov (United States)

    Abate, Davide; Saldan, Alda; Forner, Gabriella; Tinto, Daniel; Bianchin, Alice; Palù, Giorgio

    2014-02-01

    Assessing the CMV specific CMI in transplant subjects represents a promising strategy to determine the risk of infection on individual basis. In this study 61 adult CMV IgG seropositive solid organ transplant recipients were examined in order to improve the efficacy of CMI detection. For this purpose, pair-wise comparisons were conducted comparing positive control stimuli PWM and PMA/iono and CMV stimuli, pp65 peptide pool and whole CMV particle. Rosette pre-depletion of blood was also investigated for detecting CD4+ or CD8+ T-cell responses using the IFN-g ELISPOT assay. In the time-points 30-180 days after transplantation, PMA/iono produced statistically significant higher responses compared to PWM, probably because PMA/iono activation pathway is independent from the effect of immunosuppressive drugs. The data showed that 11% of transplant patients displayed very low or undetectable responses to pp65 peptide pool antigen while having sustained high responses to whole CMV particle. In addition, in all the subjects analyzed, CMI responses to CMV particle produced a statistically significant higher number of spots compared to pp65 peptide pool antigen. Rosette pre-depletion of whole blood proved to be effective in detecting CD4+ or CD8+ T-cell responses similarly to flow cytometry. Taken together, the following recommendations are suggested to optimize the CMV-ELISPOT for transplantation settings: (1) use PMA/iono as positive control; (2) whole virus particle should be used to avoid peptide-related false negative responses; (3) a rosette pre-depletion step may be useful to detect CD4+ or CD8+ T-cell responses. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Non-Tuberculous Mycobacteria and the Performance of Interferon Gamma Release Assays in Denmark

    DEFF Research Database (Denmark)

    Hermansen, Thomas Stig; Thomsen, Vibeke Østergaard; Lillebaek, Troels

    2014-01-01

    , nationwide patient data on positive NTM cultures (n = 925) were combined with nationwide data on QFT results (n = 16,133), both retrieved from the International Reference Laboratory of Mycobacteriology, Denmark. A total of 112 patients with NTM infections had a QFT performed, 53 patients had definite NTM...... disease, 10 had possible disease and 49 had NTM colonization. RESULTS: QFT was positive in 8% (4/53) of patients with definite disease, 40% (4/10) with possible disease and 31% (15/49) with colonization. Positivity rate was lowest among patients with definite disease infected with NTM without the RD1...

  19. Evaluation of immune responses in HIV infected patients with pleural tuberculosis by the QuantiFERON® TB-Gold interferon-gamma assay

    Directory of Open Access Journals (Sweden)

    Lekabe Jacob M

    2008-03-01

    Full Text Available Abstract Background Diagnosis of tuberculous (TB pleuritis is difficult and better diagnostic tools are needed. New blood based interferon-gamma (IFN-γ tests are promising, but sensitivity could be low in HIV positive patients. The IFN-γ tests have not yet been validated for use in pleural fluid, a compartment with higher level of immune activation than in blood. Methods The QuantiFERON TB®-Gold (QFT-TB test was analysed in blood and pleural fluid from 34 patients presenting with clinically suspected pleural TB. Clinical data, HIV status and CD4 cell counts were recorded. Adenosine deaminase activity (ADA analysis and TB culture were performed on pleural fluid. Results The patients were categorised as 'confirmed TB' (n = 12, 'probable TB' (n = 16 and 'non-TB' pleuritis (n = 6 based on TB culture results and clinical and biochemical criteria. The majority of the TB patients were HIV infected (82%. The QFT-TB in pleural fluid was positive in 27% and 56% of the 'confirmed TB' and 'probable TB' cases, respectively, whereas the corresponding sensitivities in blood were 58% and 83%. Indeterminate results in blood (25% were caused by low phytohemagglutinin (PHA = positive control IFN-γ responses, significantly lower in the TB patients as compared to the 'non-TB' cases (p = 0.02. Blood PHA responses correlated with CD4 cell count (r = 0.600, p = 0.028. In contrast, in pleural fluid indeterminate results (52% were caused by high Nil (negative control IFN-γ responses in both TB groups. Still, the Nil IFN-γ responses were lower than the TB antigen responses (p Conclusion The QFT-TB test in blood could contribute to the diagnosis of TB pleuritis in the HIV positive population. Still, the number of inconclusive results is too high to recommend the commercial QFT-TB test for routine use in pleural fluid in a TB/HIV endemic resource-limited setting.

  20. An Enzyme-Linked Immunosorbent Spot Assay Measuring Borrelia burgdorferi B31-Specific Interferon Gamma-Secreting T Cells Cannot Discriminate Active Lyme Neuroborreliosis from Past Lyme Borreliosis: a Prospective Study in the Netherlands.

    Science.gov (United States)

    van Gorkom, T; Sankatsing, S U C; Voet, W; Ismail, D M; Muilwijk, R H; Salomons, M; Vlaminckx, B J M; Bossink, A W J; Notermans, D W; Bouwman, J J M; Kremer, K; Thijsen, S F T

    2018-04-01

    Two-tier serology testing is most frequently used for the diagnosis of Lyme borreliosis (LB); however, a positive result is no proof of active disease. To establish a diagnosis of active LB, better diagnostics are needed. Tests investigating the cellular immune system are available, but studies evaluating the utility of these tests on well-defined patient populations are lacking. Therefore, we investigated the utility of an enzyme-linked immunosorbent spot (ELISpot) assay to diagnose active Lyme neuroborreliosis. Peripheral blood mononuclear cells (PBMCs) of various study groups were stimulated by using Borrelia burgdorferi strain B31 and various recombinant antigens, and subsequently, the number of Borrelia -specific interferon gamma (IFN-γ)-secreting T cells was measured. We included 33 active and 37 treated Lyme neuroborreliosis patients, 28 healthy individuals treated for an early manifestation of LB in the past, and 145 untreated healthy individuals. The median numbers of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 10 5 PBMCs did not differ between active Lyme neuroborreliosis patients (6.0; interquartile range [IQR], 0.5 to 14.0), treated Lyme neuroborreliosis patients (4.5; IQR, 2.0 to 18.6), and treated healthy individuals (7.4; IQR, 2.3 to 14.9) ( P = 1.000); however, the median number of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 10 5 PBMCs among untreated healthy individuals was lower (2.0; IQR, 0.5 to 3.9) ( P ≤ 0.016). We conclude that the Borrelia ELISpot assay, measuring the number of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 10 5 PBMCs, correlates with exposure to the Borrelia bacterium but cannot be used for the diagnosis of active Lyme neuroborreliosis. Copyright © 2018 van Gorkom et al.

  1. Evaluation of accuracy and uncertainty of ELISA assays for the determination of interleukin-4, interleukin-5, interferon-gamma and tumor necrosis factor-alpha

    DEFF Research Database (Denmark)

    Borg, Lone; Kristiansen, Jesper; Christensen, Jytte M

    2002-01-01

    . However, models for establishing the traceability and uncertainty of immunoassay results are lacking. Sandwich enzyme-linked immunosorbent assays (ELISAs) were developed for determination of the human cytokines interleukin-4 (IL-4), interleukin-5 (IL-5), interferon-y (IFN-gamma) and tumor necrosis factor-alpha...... (TNF-alpha). The accuracy of each of the assays was evaluated in the ranges of 1-15 microg/l (IL-4), 0.001-1 microg/l (IL-5), 0.5-2.5 microg/l (IFN-T) and 0.14-2.2 microg/l (TNF-alpha). Other evaluated performance characteristics were the limit of detection (LOD), immunological specificity......) of the assessed ELISAs was found to be in the range of 11-18%, except for IL-5 where RSDA increased at decreasing concentrations. The LOD was 0.12 microg/l, 0.0077 microg/l, 0.0069 microg/l and 0.0063 microg/l for IL-4, IL-5, IFN-gamma and TNF-alpha, respectively. Traceability to the WHO IS was established...

  2. Strong interferon-gamma mediated cellular immunity to scrub typhus demonstrated using a novel whole cell antigen ELISpot assay in rhesus macaques and humans.

    Directory of Open Access Journals (Sweden)

    Manutsanun Sumonwiriya

    2017-09-01

    Full Text Available Scrub typhus is a febrile infection caused by the obligate intracellular bacterium Orientia tsutsugamushi, which causes significant morbidity and mortality across the Asia-Pacific region. The control of this vector-borne disease is challenging due to humans being dead-end hosts, vertical maintenance of the pathogen in the vector itself, and a potentially large rodent reservoir of unclear significance, coupled with a lack of accurate diagnostic tests. Development of an effective vaccine is highly desirable. This however requires better characterization of the natural immune response of this neglected but important disease. Here we implement a novel IFN-γ ELISpot assay as a tool for studying O. tsutsugamushi induced cellular immune responses in an experimental scrub typhus rhesus macaque model and human populations. Whole cell antigen for O. tsutsugamushi (OT-WCA was prepared by heat inactivation of Karp-strain bacteria. Rhesus macaques were infected intradermally with O. tsutsugamushi. Freshly isolated peripheral blood mononuclear cells (PBMC from infected (n = 10 and uninfected animals (n = 5 were stimulated with OT-WCA, and IFN-γ secreting cells quantitated by ELISpot assay at five time points over 28 days. PBMC were then assayed from people in a scrub typhus-endemic region of Thailand (n = 105 and responses compared to those from a partially exposed population in a non-endemic region (n = 14, and to a naïve population in UK (n = 12. Mean results at Day 0 prior to O. tsutsugamushi infection were 12 (95% CI 0-25 and 15 (2-27 spot-forming cells (SFC/106 PBMC for infected and control macaques respectively. Strong O. tsutsugamushi-specific IFN-γ responses were seen post infection, with ELISpot responses 20-fold higher than baseline at Day 7 (mean 235, 95% CI 200-270 SFC/106 PBMC, 105-fold higher at Day 14 (mean 1261, 95% CI 1,097-1,425 SFC/106 PBMC, 125-fold higher at Day 21 (mean 1,498, 95% CI 1,496-1,500 SFC/106 PBMC and 118-fold higher at

  3. Strong interferon-gamma mediated cellular immunity to scrub typhus demonstrated using a novel whole cell antigen ELISpot assay in rhesus macaques and humans.

    Science.gov (United States)

    Sumonwiriya, Manutsanun; Paris, Daniel H; Sunyakumthorn, Piyanate; Anantatat, Tippawan; Jenjaroen, Kemajittra; Chumseng, Suchintana; Im-Erbsin, Rawiwan; Tanganuchitcharnchai, Ampai; Jintaworn, Suthatip; Blacksell, Stuart D; Chowdhury, Fazle R; Kronsteiner, Barbara; Teparrukkul, Prapit; Burke, Robin L; Lombardini, Eric D; Richards, Allen L; Mason, Carl J; Jones, James W; Day, Nicholas P J; Dunachie, Susanna J

    2017-09-01

    Scrub typhus is a febrile infection caused by the obligate intracellular bacterium Orientia tsutsugamushi, which causes significant morbidity and mortality across the Asia-Pacific region. The control of this vector-borne disease is challenging due to humans being dead-end hosts, vertical maintenance of the pathogen in the vector itself, and a potentially large rodent reservoir of unclear significance, coupled with a lack of accurate diagnostic tests. Development of an effective vaccine is highly desirable. This however requires better characterization of the natural immune response of this neglected but important disease. Here we implement a novel IFN-γ ELISpot assay as a tool for studying O. tsutsugamushi induced cellular immune responses in an experimental scrub typhus rhesus macaque model and human populations. Whole cell antigen for O. tsutsugamushi (OT-WCA) was prepared by heat inactivation of Karp-strain bacteria. Rhesus macaques were infected intradermally with O. tsutsugamushi. Freshly isolated peripheral blood mononuclear cells (PBMC) from infected (n = 10) and uninfected animals (n = 5) were stimulated with OT-WCA, and IFN-γ secreting cells quantitated by ELISpot assay at five time points over 28 days. PBMC were then assayed from people in a scrub typhus-endemic region of Thailand (n = 105) and responses compared to those from a partially exposed population in a non-endemic region (n = 14), and to a naïve population in UK (n = 12). Mean results at Day 0 prior to O. tsutsugamushi infection were 12 (95% CI 0-25) and 15 (2-27) spot-forming cells (SFC)/106 PBMC for infected and control macaques respectively. Strong O. tsutsugamushi-specific IFN-γ responses were seen post infection, with ELISpot responses 20-fold higher than baseline at Day 7 (mean 235, 95% CI 200-270 SFC/106 PBMC), 105-fold higher at Day 14 (mean 1261, 95% CI 1,097-1,425 SFC/106 PBMC), 125-fold higher at Day 21 (mean 1,498, 95% CI 1,496-1,500 SFC/106 PBMC) and 118-fold higher at Day 28

  4. Enzyme-linked immunospot: an alternative method for the detection of interferon gamma in Johne's disease

    DEFF Research Database (Denmark)

    Begg, Douglas J.; de Silva, Kumudika; Bosward, Katrina

    2009-01-01

    To date, the sensitivity of the interferon gamma (IFN-) enzyme-linked immunosorbent assay (ELISA) to detect Johne's disease (JD) has been poor, especially in the early stages of disease. To improve the sensitivity of IFN- detection in the early stages of infection, an alternate assay needs to be ...

  5. Atorvastatin prevents age-related and amyloid-beta-induced microglial activation by blocking interferon-gamma release from natural killer cells in the brain

    LENUS (Irish Health Repository)

    Lyons, Anthony

    2011-03-31

    Abstract Background Microglial function is modulated by several factors reflecting the numerous receptors expressed on the cell surface, however endogenous factors which contribute to the age-related increase in microglial activation remain largely unknown. One possible factor which may contribute is interferon-γ (IFNγ). IFNγ has been shown to increase in the aged brain and potently activates microglia, although its endogenous cell source in the brain remains unidentified. Methods Male Wistar rats were used to assess the effect of age and amyloid-β (Aβ) on NK cell infiltration into the brain. The effect of the anti-inflammatory compound, atorvastatin was also assessed under these conditions. We measured cytokine and chemokine (IFNγ, IL-2, monocyte chemoattractant protein-1 (MCP-1) and IFNγ-induced protein 10 kDa (IP-10)), expression in the brain by appropriate methods. We also looked at NK cell markers, CD161, NKp30 and NKp46 using flow cytometry and western blot. Results Natural killer (NK) cells are a major source of IFNγ in the periphery and here we report the presence of CD161+ NKp30+ cells and expression of CD161 and NKp46 in the brain of aged and Aβ-treated rats. Furthermore, we demonstrate that isolated CD161+ cells respond to interleukin-2 (IL-2) by releasing IFNγ. Atorvastatin, the HMG-CoA reductase inhibitor, attenuates the increase in CD161 and NKp46 observed in hippocampus of aged and Aβ-treated rats. This was paralleled by a decrease in IFNγ, markers of microglial activation and the chemokines, MCP-1 and IP-10 which are chemotactic for NK cells. Conclusions We propose that NK cells contribute to the age-related and Aβ-induced neuroinflammatory changes and demonstrate that these changes can be modulated by atorvastatin treatment.

  6. Comparison of Tuberculin Skin Test result and interferon gamma response to human PPD in BCG scar positive and negative children.

    Science.gov (United States)

    Sayyahfar, Shirin; Karimi, Abdollah; Fahimzad, Alireza; Shamshiri, Ahmad Reza

    2014-03-01

    The aim of this study is to compare Tuberculin Skin Test (TST) result and interferon gamma response to human PPD (purified protein derivative), in scar positive and scar negative BCG-vaccinated children. Between August 2007 and May 2008 a total of 236 children aged 1-168 months (mean 21 months) admitted to Mofid Children's Hospital, Tehran, Iran, were enrolled in a cross-sectional study. Each patient was examined for BCG vaccine scar and tested with TST and human PPD-based Interferon Gamma Release Assay (IGRA). Two hundred and twenty one cases out of 236 (44% female, 1-168 months, mean age 21 months) were scar positive of whom 95% TST result was negative. Human PPD-based IGRA was positive in 110 (49.8%), negative in 85 (38.4 %) and indeterminate in 26 (11.8%) of scar positive patients. Fifteen children (40% female, 1-156 months; mean age 42 months) were scar negative. All the scar negative cases were TST negative. Human PPD-based IGRA was positive in 10 (66.7%), negative in 4 (26.7%) and indeterminate in 1 (6.7%) of scar negative patients. Immune responsiveness to human PPD antigens in scar positive and negative children may not correspond with results of the Tuberculin Skin Test. Copyright © 2013 Ministry of Health, Saudi Arabia. Published by Elsevier Ltd. All rights reserved.

  7. Interferon-gamma response to the treatment of active pulmonary and extra-pulmonary tuberculosis.

    Science.gov (United States)

    Liang, L; Shi, R; Liu, X; Yuan, X; Zheng, S; Zhang, G; Wang, W; Wang, J; England, K; Via, L E; Cai, Y; Goldfeder, L C; Dodd, L E; Barry, C E; Chen, R Y

    2017-10-01

    Interferon-gamma (IFN-γ) release assays (IGRAs) are used to diagnose tuberculosis (TB) but not to measure treatment response. To measure IFN-γ response to active anti-tuberculosis treatment. Patients from the Henan Provincial Chest Hospital, Henan, China, with TB symptoms and/or signs were enrolled into this prospective, observational cohort study and followed for 6 months of treatment, with blood and sputum samples collected at 0, 2, 4, 6, 8, 16 and 24 weeks. The QuantiFERON® TB-Gold assay was run on collected blood samples. Participants received a follow-up telephone call at 24 months to determine relapse status. Of the 152 TB patients enrolled, 135 were eligible for this analysis: 118 pulmonary (PTB) and 17 extra-pulmonary TB (EPTB) patients. IFN-γ levels declined significantly over time among all patients (P = 0.002), with this decline driven by PTB patients (P = 0.001), largely during the initial 8 weeks of treatment (P = 0.019). IFN-γ levels did not change among EPTB patients over time or against baseline culture or drug resistance status. After 6 months of effective anti-tuberculosis treatment, IFN-γ levels decreased significantly in PTB patients, largely over the initial 8 weeks of treatment. IFN-γ concentrations may offer some value for monitoring anti-tuberculosis treatment response among PTB patients.

  8. Pulmonary Immune-Compartment-Specific Interferon Gamma Responses in HIV-Infected Individuals with Active Tuberculosis (TB in an Area of High TB Prevalence

    Directory of Open Access Journals (Sweden)

    S. Buldeo

    2012-01-01

    Full Text Available There is a paucity of data on the pulmonary immune-compartment interferon gamma (IFNγ response to M. tuberculosis, particularly in settings of high tuberculosis (TB prevalence and in HIV-coinfected individuals. This data is necessary to understand the diagnostic potential of commercially available interferon gamma release assays (IGRAs in both the pulmonary immune-compartment and peripheral blood. We used intracellular cytokine staining by flow cytometry to assess the IFNγ response to purified protein derivative (PPD and early secretory antigen 6 (ESAT6 in induced sputa (ISp and blood samples from HIV-infected, smear-negative, TB suspects. We found that individuals with active TB disease produced significantly less IFNγ in response to PPD in their induced sputa samples than individuals with non-active TB (control group. This difference was not reflected in the peripheral blood, even within the CD27− CD4+ memory T lymphocyte population. These findings suggest that progression to active TB disease may be associated with the loss of IFNγ secretion at the site of primary infection. Our findings highlight the importance of studying pulmonary immune-compartment M. tuberculosis specific responses to elucidate IFNγ secretion across the spectrum of TB disease.

  9. Interferon gamma peptidomimetic targeted to interstitial myofibroblasts attenuates renal fibrosis after unilateral ureteral obstruction in mice

    NARCIS (Netherlands)

    Poosti, Fariba; Bansal, Ruchi; Yazdani, Saleh; Prakash, Jai; Beljaars, Leonie; van den Born, Jacob; de Borst, Martin H.; van Goor, Harry; Hillebrands, Jan-Luuk; Poelstra, Klaas

    2016-01-01

    Renal fibrosis cannot be adequately treated since anti-fibrotic treatment is lacking. Interferon-gamma is a pro-inflammatory cytokine with anti-fibrotic properties. Clinical use of interferon-gamma is hampered due to inflammation-mediated systemic side effects. We used an interferon-gamma

  10. Tuberculin skin test and QuantiFERON® assay in young children investigated for tuberculosis in South Africa

    NARCIS (Netherlands)

    Moyo, S.; Isaacs, F.; Gelderbloem, S.; Verver, S.; Hawkridge, A. J.; Hatherill, M.; Tameris, M.; Geldenhuys, H.; Workman, L.; Pai, M.; Hussey, G.; Hanekom, W. A.; Mahomed, H.

    2011-01-01

    Although the literature on interferon-gamma release assays on tuberculosis (TB) in children has increased, data pertaining to young children remain relatively limited. To compare results from the tuberculin skin test (TST) and the QuantiFERON®-TB Gold In-Tube assay (QFT) in children aged <3 years

  11. Household food insecurity is associated with low interferon-gamma levels in pregnant Indian women.

    Science.gov (United States)

    Vaidya, A; Bhosale, R; Sambarey, P; Suryavanshi, N; Young, S; Mave, V; Kanade, S; Kulkarni, V; Deshpande, P; Balasubramanian, U; Elf, J; Gupte, N; Gupta, A; Mathad, J S

    2017-07-01

    Over 20% of tuberculosis (TB) cases during pregnancy occur in India. To determine the association between household food insecurity and interferon-gamma (IFN-γ) levels in pregnancy. Pregnant women in India were administered the Household Food Insecurity Access Scale (HFIAS) questionnaire and underwent an IFN-γ release assay. Logistic regression was used to identify factors associated with food insecurity. Of 538 women, 60 (11%) had household food insecurity, 47 (78%) of which were moderate or severe food insecure. After mitogen stimulation, moderate or severe food insecure women had a median IFN-γ concentration of 4.2 IU/ml (IQR 2.2-9.8) vs. 8.4 IU/ml (IQR 3.0-10) in women with no or mild food insecurity (P = 0.03). In multivariate analysis, higher IFN-γ concentrations were associated with human immunodeficiency virus infection (OR 1.3, 95%CI 0.51-2.1, P = 0.001), and inversely associated with moderate or severe food insecurity (OR -1.6, 95%CI -2.9 to -0.27, P = 0.02) and the number of adults in the household (OR -0.08, 95%CI -0.16 to -0.01, P = 0.03). There was no association between food insecurity and IFN-γ response to Mycobacterium tuberculosis antigen. Food insecurity in pregnancy is associated with low IFN-γ levels. There was no association between food insecurity and IFN-γ response to M. tuberculosis antigen, but our study was underpowered to detect this outcome.

  12. Tuberculin skin test and interferon-gamma release assay values are associated with antimicrobial peptides expression in  polymorphonuclear cells during latent tuberculous infection

    Directory of Open Access Journals (Sweden)

    Julio E Castañeda-Delgado

    2014-06-01

    Full Text Available It has been reported that patients with progressive tuberculosis (TB express abundant amounts of the antimicrobial peptides (AMPs cathelicidin (LL-37 and human neutrophil peptide-1 (HNP-1 in circulating cells, whereas latent TB infected donors showed no differences when compared with purified protein derivative (PPD and QuantiFERON®-TB Gold (QFT-healthy individuals. The aim of this study was to determine whether LL-37 and HNP-1 production correlates with higher tuberculin skin test (TST and QFT values in TB household contacts. Twenty-six TB household contact individuals between 26-58 years old TST and QFT positive with at last two years of latent TB infection were recruited. AMPs production by polymorphonuclear cells was determined by flow cytometry and correlation between TST and QFT values was analysed. Our results showed that there is a positive correlation between levels of HNP-1 and LL-37 production with reactivity to TST and/or QFT levels. This preliminary study suggests the potential use of the expression levels of these peptides as biomarkers for progression in latent infected individuals.

  13. Consistency of Mycobacterium tuberculosis-Specific Interferon-Gamma Responses in HIV-1-Infected Women during Pregnancy and Postpartum

    Directory of Open Access Journals (Sweden)

    Sasi R. Jonnalagadda

    2012-01-01

    Full Text Available Background. We determined the consistency of positive interferon-gamma (IFN-γ release assays (IGRAs to detect latent TB infection (LTBI over one-year postpartum in HIV-1-infected women. Methods. Women with positive IGRAs during pregnancy had four 3-monthly postpartum IGRAs. Postpartum change in magnitude of IFN-γ response was determined using linear mixed models. Results. Among 18 women with positive pregnancy IGRA, 15 (83% had a subsequent positive IGRA; 9 (50% were always positive, 3 (17% were always negative, and 6 (33% fluctuated between positive and negative IGRAs. Women with pregnancy IGRA IFN-γ >8 spot forming cells (SFCs/well were more likely to have consistent postpartum IGRA response (odds ratio: 10.0; 95% confidence interval (CI: 0.9–117.0. Change in IFN-γ response over postpartum was 10.2 SFCs/well (95% CI: −1.5–21.8 SFCs/well. Conclusion. Pregnancy positive IGRAs were often maintained postpartum with increased consistency in women with higher baseline responses. There were modest increases in magnitude of IGRA responses postpartum.

  14. Lactobacillus GG has in vitro effects on enhanced interleukin-10 and interferon-gamma release of mononuclear cells but no in vivo effects in supplemented mothers and their neonates.

    Science.gov (United States)

    Kopp, M V; Goldstein, M; Dietschek, A; Sofke, J; Heinzmann, A; Urbanek, R

    2008-04-01

    The value of probiotics for primary prevention is controversial. Moreover, only little is known about the underlying immunological mechanisms of action. Therefore, we assessed the proliferative response and cytokine release in cultures of isolated mononuclear cells from pregnant women and their neonates supplemented with Lactobacillus GG (LGG) or placebo. In a double-blind, placebo-controlled prospective trial, pregnant women with at least one first-degree relative or a partner with an atopic disease were randomly assigned to receive either the probiotic LGG (ATCC 53103; 5 x 10(9) colony-forming units LGG twice daily) or placebo 4-6 weeks before expected delivery, followed by a post-natal period of 6 months. Cord blood mononuclear cells (CBMC) and peripheral blood mononuclear cells (PBMC) of the corresponding mother were isolated from cord blood and peripheral blood (n=68). The proliferative response of CBMC and PBMC was expressed as the stimulation index (SI), which was calculated according to the ratio between the mean counts per minute (c.p.m.) values measured in the wells with stimulated cells and the mean c.p.m. values measured in the wells with unstimulated cells. Additionally, the cytokines IFN-gamma, IL-10 and IL-13 in the cell culture supernatants were measured using the ELISA technique. No difference was observed between the LGG-supplemented group and the placebo group in terms of the proliferative capacity of maternal or neonatal cord blood cells in response to IL-2, beta-lactoglobulin or LGG. In vitro stimulation with LGG resulted in significantly enhanced release of IL-10 and IFN-gamma, compared with cytokine release in unstimulated controls. However, this phenomenon was observed in supernatants of maternal and neonatal MC in both groups, independent of prior supplementation with LGG. LGG has in vitro effects on enhanced IL-10 and IFN-gamma release of mononuclear cells. However, supplementation with LGG during pregnancy did not alter the proliferative

  15. Specificity of the Tuberculin Skin Test and the T-SPOT."TB" Assay among Students in a Low-Tuberculosis Incidence Setting

    Science.gov (United States)

    Talbot, Elizabeth A.; Harland, Dawn; Wieland-Alter, Wendy; Burrer, Sherry; Adams, Lisa V.

    2012-01-01

    Objective: Interferon-[gamma] release assays (IGRAs) are an important tool for detecting latent "Mycobacterium tuberculosis" infection (LTBI). Insufficient data exist about IGRA specificity in college health centers, most of which screen students for LTBI using the tuberculin skin test (TST). Participants: Students at a low-TB incidence college…

  16. Interferon Gamma in African Trypanosome Infections: Friends or Foes?

    Science.gov (United States)

    Wu, Hui; Liu, Gongguan; Shi, Meiqing

    2017-01-01

    African trypanosomes cause fatal infections in both humans and livestock. Interferon gamma (IFN-γ) plays an essential role in resistance to African trypanosomes. However, increasing evidence suggests that IFN-γ, when excessively synthesized, also induces immunopathology, enhancing susceptibility to the infection. Thus, production of IFN-γ must be tightly regulated during infections with African trypanosomes to ensure that a robust immune response is elicited without tissue destruction. Early studies have shown that secretion of IFN-γ is downregulated by interleukin 10 (IL-10). More recently, IL-27 has been identified as a negative regulator of IFN-γ production during African trypanosome infections. In this review, we discuss the current state of our understanding of the role of IFN-γ in African trypanosome infections. We have focused on the cellular source of IFN-γ, its beneficial and detrimental effects, and mechanisms involved in regulation of its production, highlighting some recent advances and offering some perspectives on future directions.

  17. Evaluation of Interferon-gamma Application for Recognition of Patients Afflicted by Non-healing Cutaneous Leishmaniasis

    OpenAIRE

    Mohammad Moafi; Hossein Rezvan; Roya Sherkat; Sayyed Hamid Zarkesh-Esfahani; Roya Taleban; Ali Asilian; Mohammad Ali Nilforoushzadeh; Fariba Jaffary; Marjan Mansourian; Fatemeh Sokhanvari; Nazli Ansari

    2017-01-01

    Introduction:Different studies undertaken in the animal modeling show that Interferon-gamma deficiency impairs healing process of Leishmania infection. It seems that the level of Interferon-gamma production could also affect the healing duration of Leishmania lesion in humans. The current study aims to investigate the possibility of Interferon-gamma application for recognition of cases afflicted by non-healing Leishmaniasis. Materials and methods: Peripheral blood mononuclear cells (PBMCs) of...

  18. MOLECULAR CLONING, SEQUENCING, EXPRESSION AND BIOLOGICAL ACTIVITY OF GIANT PANDA (AILUROPODA MELANOLEUCA) INTERFERON-GAMMA.

    Science.gov (United States)

    Zhu, Hui; Wang, Wen-Xiu; Wang, Bao-Qin; Zhu, Xiao-Fu; Wu, Xu-Jin; Ma, Qing-Yi; Chen, De-Kun

    2012-06-29

    The giant panda (Ailuropoda melanoleuca) is an endangered species and indigenous to China. Interferon-gamma (IFN-γ) is the only member of type □ IFN and is vital for the regulation of host adapted immunity and inflammatory response. Little is known aboutthe FN-γ gene and its roles in giant panda.In this study, IFN-γ gene of Qinling giant panda was amplified from total blood RNA by RT-CPR, cloned, sequenced and analysed. The open reading frame (ORF) of Qinling giant panda IFN-γ encodes 152 amino acidsand is highly similar to Sichuan giant panda with an identity of 99.3% in cDNA sequence. The IFN-γ cDNA sequence was ligated to the pET32a vector and transformed into E. coli BL21 competent cells. Expression of recombinant IFN-γ protein of Qinling giant panda in E. coli was confirmed by SDS-PAGE and Western blot analysis. Biological activity assay indicated that the recombinant IFN-γ protein at the concentration of 4-10 µg/ml activated the giant panda peripheral blood lymphocytes,while at 12 µg/mlinhibited. the activation of the lymphocytes.These findings provide insights into the evolution of giant panda IFN-γ and information regarding amino acid residues essential for their biological activity.

  19. Association between level of interferon gamma and acid-fast bacillipositivity in pulmonary tuberculosis

    Science.gov (United States)

    Priwahyuningtyas, N. B.; Sinaga, B. Y. M.; Pandia, P.; Eyanoer, P. C.

    2018-03-01

    Tuberculosis is an infectious disease which caused by Mycobacterium tuberculosis (M. tuberculosis) that infected numerous organ especially the lung. A person’s immunity is very affecting for a person exposed to pulmonary tuberculosis. T-helper-1 cell (Th1) is very influential in the immune system especially in interfering intracellular bacterial infection. One of the cytokines known produced by Th1 cell is interferon gamma (IFN-γ) which is in eliminating M. tuberculosis. The study aims to identify the association between level of IFN-γ and AFB positivity in pulmonary tuberculosis patients in Medan. It is a case-control study. The subjects of the study were 60 new cases of pulmonary tuberculosis with AFB sputum smear- positive that never received ATT consisting 20 cases AFB (+1), 20 cases AFB (+2) and 20 cases AFB (+3).Samples were plasma collected from the venous blood of pulmonary tuberculosis patients. The plasma then underwent laboratory assay with ELISA techniques. Independent t-test was p<0.05 considered significant. Level of IFN-γ in TB AFB (+1) is higher than TB AFB (+2) and (+3), with thesignificant statistical result (p=0.001).

  20. Whole-exome sequencing reveals a rare interferon gamma receptor 1 mutation associated with myasthenia gravis.

    Science.gov (United States)

    Qi, Guoyan; Liu, Peng; Gu, Shanshan; Yang, Hongxia; Dong, Huimin; Xue, Yinping

    2018-04-01

    Our study is aimed to explore the underlying genetic basis of myasthenia gravis. We collected a Chinese pedigree with myasthenia gravis, and whole-exome sequencing was performed on the two affected siblings and their parents. The candidate pathogenic gene was identified by bioinformatics filtering, which was further verified by Sanger sequencing. The homozygous mutation c.G40A (p.V14M) in interferon gamma receptor 1was identified. Moreover, the mutation was also detected in 3 cases of 44 sporadic myasthenia gravis patients. The p.V14M substitution in interferon gamma receptor 1 may affect the signal peptide function and the translocation on cell membrane, which could disrupt the binding of the ligand of interferon gamma and antibody production, contributing to myasthenia gravis susceptibility. We discovered that a rare variant c.G40A in interferon gamma receptor 1 potentially contributes to the myasthenia gravis pathogenesis. Further functional studies are needed to confirm the effect of the interferon gamma receptor 1 on the myasthenia gravis phenotype.

  1. Interferon gamma peptidomimetic targeted to hepatic stellate cells ameliorates acute and chronic liver fibrosis in vivo

    NARCIS (Netherlands)

    Bansal, Ruchi; Prakash, Jai; De Ruiter, Marieke; Poelstra, Klaas

    2014-01-01

    Hepatic stellate cells play a crucial role in the pathogenesis of hepatic fibrosis. Thus, pharmacological inhibition of pro-fibrotic activities of these cells might lead to an effective therapy for this disease. Among the potent antifibrotics, interferon gamma (IFN gamma), a proinflammatory

  2. Interferon gamma peptidomimetic targeted to hepatic stellate cells ameliorates acute and chronic liver fibrosis in vivo

    NARCIS (Netherlands)

    Bansal, Ruchi; Prakash, Jai; de Ruiter, Marieke; Poelstra, Klaas

    2014-01-01

    Hepatic stellate cells play a crucial role in the pathogenesis of hepatic fibrosis. Thus, pharmacological inhibition of pro-fibrotic activities of these cells might lead to an effective therapy for this disease. Among the potent anti-fibrotics, interferon gamma (IFNγ), a proinflammatory cytokine, is

  3. Review of the recombinant human interferon gamma as an immunotherapeutic: Impacts of production platforms and glycosylation.

    Science.gov (United States)

    Razaghi, Ali; Owens, Leigh; Heimann, Kirsten

    2016-12-20

    Human interferon gamma is a cytokine belonging to a diverse group of interferons which have a crucial immunological function against mycobacteria and a wide variety of viral infections. To date, it has been approved for treatment of chronic granulomatous disease and malignant osteopetrosis, and its application as an immunotherapeutic agent against cancer is an increasing prospect. Recombinant human interferon gamma, as a lucrative biopharmaceutical, has been engineered in different expression systems including prokaryotic, protozoan, fungal (yeasts), plant, insect and mammalian cells. Human interferon gamma is commonly expressed in Escherichia coli, marketed as ACTIMMUNE ® , however, the resulting product of the prokaryotic expression system is unglycosylated with a short half-life in the bloodstream; the purification process is tedious and makes the product costlier. Other expression systems also did not show satisfactory results in terms of yields, the biological activity of the protein or economic viability. Thus, the review aims to synthesise available information from previous studies on the production of human interferon gamma and its glycosylation patterns in different expression systems, to provide direction to future research in this field. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Crystal structure of human interferon-gamma receptor 2 reveals the structural basis for receptor specificity

    Czech Academy of Sciences Publication Activity Database

    Mikulecký, Pavel; Zahradník, Jiří; Kolenko, Petr; Černý, Jiří; Charnavets, Tatsiana; Kolářová, Lucie; Nečasová, Iva; Pham, Phuong Ngoc; Schneider, Bohdan

    2016-01-01

    Roč. 72, č. 9 (2016), s. 1017-1025 ISSN 2059-7983 R&D Projects: GA ČR(CZ) GA16-20507S; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:86652036 Keywords : interferon-gamma receptor 2 * fibronectin type III domain * class 2 cytokine receptors Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.114, year: 2016

  5. Evaluation of Interferon-gamma Application for Recognition of Patients Afflicted by Non-healing Cutaneous Leishmaniasis

    Directory of Open Access Journals (Sweden)

    Mohammad Moafi

    2017-06-01

    Full Text Available Introduction:Different studies undertaken in the animal modeling show that Interferon-gamma deficiency impairs healing process of Leishmania infection. It seems that the level of Interferon-gamma production could also affect the healing duration of Leishmania lesion in humans. The current study aims to investigate the possibility of Interferon-gamma application for recognition of cases afflicted by non-healing Leishmaniasis. Materials and methods: Peripheral blood mononuclear cells (PBMCs of 32 patients, who were afflicted by healing or non-healing Leishmaniasis, were isolated and the levels of interferon-gamma were determined, using ELISA method. Afterwards, the cut-off point of interferon-gamma to identify patients afflicted by non-healing Leishmaniasis was calculated through ROC-Curve analysis. Furthermore, Leishmanin Skin Test (LST was performed for every patient. Results: Levels of Interferon-gamma produced by PBMCs stimulated with Soluble Leishmania Antigen (SLA or Phytohemaglotinine were significantly higher in healing patients, compared with non-healing individuals (p

  6. In vitro immunomodulation of a whole blood IFN-γ release assay enhances T cell responses in subjects with latent tuberculosis infection.

    Directory of Open Access Journals (Sweden)

    Rajiv L Gaur

    Full Text Available Activation of innate immunity via pathogen recognition receptors (PRR modulates adaptive immune responses. PRR ligands are being exploited as vaccine adjuvants and as therapeutics, but their utility in diagnostics has not been explored. Interferon-gamma (IFN-γ release assays (IGRAs are functional T cell assays used to diagnose latent tuberculosis infection (LTBI; however, novel approaches are needed to improve their sensitivity.In vitro immunomodulation of a whole blood IGRA (QuantiFERON®-TB GOLD In-Tube with Toll-like receptor agonists poly(I:C, LPS, and imiquimod was performed on blood from subjects with LTBI and negative controls.In vitro immunomodulation significantly enhanced the response of T cells stimulated with M. tuberculosis antigens from subjects with LTBI but not from uninfected controls. Immunomodulation of IGRA revealed T cell responses in subjects with LTBI whose T cells otherwise do not respond to in vitro stimulation with antigens alone. Similar to their in vivo functions, addition of poly(I:C and LPS to whole blood induced secretion of inflammatory cytokines and IFN-α and enhanced the surface expression of antigen presenting and costimulatory molecules on antigen presenting cells.In vitro immunomodulation of whole blood IGRA may be an effective strategy for enhancing the sensitivity of T cells for diagnosis of LTBI.

  7. The Effect of Levothyroxine on Serum Levels of Interleukin 10 and Interferon-gamma in Rat Model of Multiple Sclerosis

    Directory of Open Access Journals (Sweden)

    Cobra Payghani

    2017-01-01

    Full Text Available Background: There is an increase in inflammatory and a reduction in anti-inflammatory cytokines in multiple sclerosis (MS. Considering the role of thyroid hormones in the development and regulation of both neural and immune systems, the aim of this study was to evaluate the effects of levothyroxine on serum concentrations of interleukin-10 (IL-10 and interferon gamma (IFN-γ in animal models of MS. Materials and Methods: To induce demyelination in male Wistar rats, lysolecithin was injected into the optic chiasm. Then levothyroxine was injected intraperitoneally (20, 50, and 100 μg/kg for 21 days. Serum levels of cytokines were measured by enzyme-linked immunosorbent assay at 7, 14, and 21 days after that. Results: The results showed that injection of lysolecithin to the optic chiasm only increased serum concentrations of IL-10 compared to the sham group (P < 0.05 at 7th day, but this increase was prevented by all doses of levothyroxine. IFN-γ was decreased significantly (P < 0.001 21 days after. Comparing to the sham group at all sampling time and with respect to the MS group at the days 7 and 21, levothyroxine decreased serum concentrations of IFN-γ significantly. Conclusion: The results showed that thyroid hormones probably could produce protective effects against induced demyelination through affecting immune responses.

  8. Evaluation of pre- and posttransplantation serum interferon-gamma and soluble CD30 for predicting liver allograft rejection.

    Science.gov (United States)

    Kim, K H; Oh, E-J; Jung, E-S; Park, Y-J; Choi, J Y; Kim, D-G; Lee, K Y; Kang, C S

    2006-06-01

    The aim of the present study was to identify whether the serum interferon-gamma (IFNgamma), a Th1 cytokine, or soluble CD30 (sCD30), a marker for activation of Th2 cytokine-producing T cells, predict acute cellular rejection episodes among liver graft patients. Pretransplant and posttransplant sera from 32 living donor liver transplant recipients obtained on days 1, 3, and 7 after surgery were tested for serum IFNgamma and sCD30 concentrations using commercial enzyme-linked immunosorbent assay kits. Recipients with an acute rejection episode (ARE) (n=14) displayed significantly higher IFNgamma concentrations pretransplant than did the patients with no ARE (n=18) (PsCD30 were not different between the non-ARE and ARE groups. However, in comparison with the non-ARE group, who showed steadily decreasing serum sCD30 levels after transplantation, 12 among the 14 patients in the ARE group showed increasing sCD30 levels from day 1 to day 3 after transplantation (PsCD30 increment during the early period after liver transplantation affects the immune response of rejection. This observation emphasizes the clinical relevance of serum sCD30, in addition to serum IFNgamma, as predictive markers for acute liver graft rejection.

  9. Equine interferon gamma synthesis in lymphocytes after in vivo infection and in vitro stimulation with EHV-1.

    Science.gov (United States)

    Paillot, R; Daly, J M; Juillard, V; Minke, J M; Hannant, D; Kydd, J H

    2005-08-22

    Equine cytotoxic T lymphocyte (CTL) responses to equine herpesvirus-1 (EHV-1) are well characterised but little is known about the cytokine response after infection or vaccination. EHV-1 is common in horses and infects lymphocytes in vivo. This virus was used as a model to measure the synthesis of interferon gamma (IFN-gamma) by equine peripheral blood mononuclear cells (PBMC) after in vivo infection and/or in vitro stimulation with EHV-1. Both flow cytometry and ELISPOT assays were used to quantify equine IFN-gamma using a mouse anti-bovine IFN-gamma monoclonal antibody (clone CC302; shown to cross-react with recombinant equine IFN-gamma) and a rabbit anti-canine IFN-gamma polyclonal antibody. The percentage of PBMC synthesising IFN-gamma after in vitro stimulation with EHV-1 increased with age. In yearlings infected experimentally with EHV-1, PBMC showed two peaks of IFN-gamma synthesis, 11 and 56 days after infection. The IFN-gamma synthesis was principally associated with CD8(+) cells. The patterns of IFN-gamma synthesis detected by intracellular IFN-gamma staining or ELISPOT were compared with CTL data and shown to be similar. These methods were also applied successfully to frozen samples of PBMC. Measurement of equine IFN-gamma using these simple techniques can now be applied to future studies on protective cellular immune responses following virus infection and/or vaccination of horses.

  10. Molecular characterisation and expression analysis of interferon gamma in response to natural Chlamydia infection in the koala, Phascolarctos cinereus.

    Science.gov (United States)

    Mathew, Marina; Pavasovic, Ana; Prentis, Peter J; Beagley, Kenneth W; Timms, Peter; Polkinghorne, Adam

    2013-09-25

    Interferon gamma (IFNγ) is a key Th1 cytokine, with a principal role in the immune response against intracellular organisms such as Chlamydia. Along with being responsible for significant morbidity in human populations, Chlamydia is also responsible for wide spread infection and disease in many animal hosts, with reports that many Australian koala subpopulations are endemically infected. An understanding of the role played by IFNγ in koala chlamydial diseases is important for the establishment of better prophylactic and therapeutic approaches against chlamydial infection in this host. A limited number of IFNγ sequences have been published from marsupials and no immune reagents to measure expression have been developed. Through preliminary analysis of the koala transcriptome, we have identified the full coding sequence of the koala IFNγ gene. Transcripts were identified in spleen and lymph node tissue samples. Phylogenetic analysis demonstrated that koala IFNγ is closely related to other marsupial IFNγ sequences and more distantly related to eutherian mammals. To begin to characterise the role of this important cytokine in the koala's response to chlamydial infection, we developed a quantitative real time PCR assay and applied it to a small cohort of koalas with and without active chlamydial disease, revealing significant differences in expression patterns between the groups. Description of the IFNγ sequence from the koala will not only assist in understanding this species' response to its most important pathogen but will also provide further insight into the evolution of the marsupial immune system. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Interferon-gamma in progression to chronic demyelination and neurological deficit following acute EAE

    DEFF Research Database (Denmark)

    Renno, T; Taupin, V; Bourbonnière, L

    1998-01-01

    The cytokine interferon-gamma (IFNgamma) is implicated in the induction of acute CNS inflammation, but it is less clear what role if any IFNgamma plays in progression to chronic demyelination and neurological deficit. To address this issue, we have expressed IFNgamma in myelinating oligodendrocytes....... In contrast to control mice, which remit from EAE with resolution of glial reactivity and leukocytic infiltration, transgenics showed chronic neurological deficits. While activated microglia/macrophages persisted in demyelinating lesions for over 100 days, CD4(+) T lymphocytes were no longer present in CNS...

  12. Effects of interferon-gamma and tumor necrosis factor-alpha on macrophage enzyme levels

    Science.gov (United States)

    Pierangeli, Silvia S.; Sonnenfeld, Gerald

    1989-01-01

    Murine peritoneal macrophages were treated with interferon-gamma (IFN-gamma) or tumor necrosis factor-alpha (TNF). Measurements of changes in acid phosphatase and beta-glucuronidase levels were made as an indication of activation by cytokine treatment. IFN-gamma or TNF-gamma treatment resulted in a significant increase in the activities of both enzymes measured in the cell lysates. This increase was observable after 6 h of incubation, but reached its maximum level after 24 h of incubation. The effect of the treatment of the cell with both cytokines together was additive. No synergistic effect of addition of both cytokines on the enzyme levels was observed.

  13. Nitric oxide selectively decreases interferon-gamma expression by activated human T lymphocytes via a cGMP-independent mechanism

    NARCIS (Netherlands)

    Roozendaal, R; Vellenga, E; Postma, DS; De Monchy, JGR; Kauffman, HF

    1999-01-01

    The role of exogenous nitric oxide (NO) on the expression of interleukin (IL)-2, IL-4, IL-5 and interferon-gamma (IFN-gamma) by freshly isolated human T lymphocytes was investigated. The presence of NO, generated from any of the NO-donor compounds, S-nitroso-N-acetyl-D,L-penicillamine (NAP),

  14. Interferon-gamma (IFN-gamma) treatment decreases the inflammatory response in chronic Pseudomonas aeruginosa pneumonia in rats

    DEFF Research Database (Denmark)

    Johansen, H K; Hougen, H P; Rygaard, J

    1996-01-01

    In a rat model of chronic Pseudomonas aeruginosa lung infection mimicking cystic fibrosis (CF), we studied whether the inflammatory response could be altered by intraperitoneal treatment with recombinant rat interferon-gamma (rrIFN-gamma). Rats were treated either before or after intratracheal ch...

  15. Surface plasmon resonance biosensor based on engineered proteins for direct detection of interferon-gamma in diluted blood plasma

    Czech Academy of Sciences Publication Activity Database

    Šípová, Hana; Ševců, Veronika; Kuchař, Milan; Ahmad, Jawid Nazir; Mikulecký, Pavel; Osičková, Adriana; Malý, Petr; Homola, Jiří

    2012-01-01

    Roč. 174, č. 11 (2012), s. 306-311 ISSN 0925-4005 R&D Projects: GA AV ČR KAN200670701 Institutional support: RVO:67985882 ; RVO:61388971 ; RVO:86652036 Keywords : Interferon gamma * Surface plasmon resonance * Biosensor Subject RIV: JB - Sensors, Measurment, Regulation Impact factor: 3.535, year: 2012

  16. Interleukin-15 differentially enhances the expression of interferon-gamma and interleukin-4 in activated human (CD4(+))T lymphocytes

    NARCIS (Netherlands)

    Borger, P; Kauffman, HF; Postma, DS; Esselink, MT; Vellenga, E

    In this study interleukin (IL)-15 was examined for its ability to modulate the expression of interferon-gamma (IFN-gamma) and IL-4 in activated human T lymphocytes. The effect of IL-15 was compared with IL-2 and IL-7, cytokines all known to use the IL-2 receptor gamma(C) chain. The results

  17. The immunomodulatory effects of interferon-gamma on mature B-lymphocyte responses.

    Science.gov (United States)

    Jurado, A; Carballido, J; Griffel, H; Hochkeppel, H K; Wetzel, G D

    1989-06-15

    Interferon-gamma (IFN-gamma) exerts a broad spectrum of activities which affect the responses of mature B-cells. It strongly inhibits B-cell activation, acts as a B-cell growth factor (BCGF), and also induces final differentiation to immunoglobulin (Ig) production. IFN-gamma is deeply involved in the differential control of isotype expression, as it enhances IgG2a production and suppresses both IgG1 and IgE production. Although it is now possible to draw a general scheme of the effects of IFN-gamma on B-cells, a number of paradoxical results still exist in the field. In this manuscript, different experimental systems are analyzed in an attempt to explain these apparent paradoxes.

  18. Interferon gamma increases survival in urine experimental cryptococcosis El Interferon gamma incrementa la sobrevida de un modelo experimental murino de criptococosis

    Directory of Open Access Journals (Sweden)

    Amadeo J. Bava

    1995-10-01

    Full Text Available Systemic disease by Cryptococcus neoformans (C. neoformans is a common opportunistic infection in immunodeficient patients. Cellular immunity seems to be the most important determinant of resistance. The aim of this study was to assess the effect of recombinant rat interferon gamma (IFN-gamma in murine cryptococcosis (Balb/c mice infected by IP route with the Rivas strain of C. neoformans, evaluating survival time, macroscopic and microscopic examination of the organs, and massive seeding of brain homogenate. IFN-gamma treatment, at a daily dose of 10,000 IU, did not modify significantly these variables when mice were challenged with a high inoculum (10(7 yeasts and treatment was delayed to 5 days after infection (median survival 21 days in control mice vs. 23 days in IFN-treated. Another set of experiments suggested that IFN-gamma treatment, at a dose of 10,000 IU/day, begun at the moment of infection could be useful (it prolonged survival from 20 to 28 days, although the difference did not achieve statistical signification. When used simultaneously with infection by 3.5 x 10(5 yeasts, IFN-gamma at 10,000 IU/day for 15 days significantly prolonged survival of mice (p = 0.004. These results suggest that, depending on the experimental conditions, IFN-gamma can improve survival of mice infected with a lethal dose of C. neoformans.Se evaluó la efectividad del interferon-gamma (IFN-gamma recombinante de rata en un modelo experimental de criptococosis desarollado en ratones Balb/C inoculados por vía intraperitoneal con la cepa Rivas de Cryptococcus neoformans (C. neoformans. Se tuvieron en cuenta el tiempo de sobrevida de los animales, el aspecto macroscópico de los órganos en la autopsia, la presencia de levaduras capsuladas en los tejidos y la siembra masiva de un homogenato de cerebro. El tratamiento con IFN-gamma, en dosis diarias de 10.000 UI, no modificó estos parámetros cuando la dosis infectante fue de 10(7 levaduras y el tratamiento se

  19. Comparison of two interferon-gamma assays and tuberculin skin test for tracing tuberculosis contacts

    NARCIS (Netherlands)

    Arend, Sandra M.; Thijsen, Steven F. T.; Leyten, Eliane M. S.; Bouwman, John J. M.; Franken, Willeke P. J.; Koster, Ben F. P. J.; Cobelens, Frank G. J.; van Houte, Arend-Jan; Bossink, Ailko W. J.

    2007-01-01

    The tuberculin skin test (TST) has low specificity. QuantiFERON-TB Gold (QFT-G) and T-SPOT.TB are based on interferon (IFN)-gamma responses to Mycobacterium tuberculosis-specific antigens. A novel in-tube format of QFT-G (QFT-GIT) offers logistical advantages. To compare TST, QFT-GIT, and T-SPOT.TB

  20. The elephant interferon gamma assay: a contribution to diagnosis of tuberculosis in elephants

    NARCIS (Netherlands)

    Angkawanish, T.; Morar, D.; Kooten, P.J.; Bontekoning, I.; Schreuder, J.; Maas, M.; Wajjwalku, W.; Sirimalaisuwan, A.; Michel, A.L.; Tijhaar, E.; Rutten, V.P.M.G.

    2013-01-01

    Mycobacterium tuberculosis (M. tb) has been shown to be the main causative agent of tuberculosis in elephants worldwide. M. tb may be transmitted from infected humans to other species including elephants and vice versa, in case of prolonged intensive contact. An accurate diagnostic approach covering

  1. Temporal Regulation of Natural Killer T Cell Interferon Gamma Responses by β-Catenin-Dependent and -Independent Wnt Signaling.

    Science.gov (United States)

    Kling, Jessica C; Jordan, Margaret A; Pitt, Lauren A; Meiners, Jana; Thanh-Tran, Thao; Tran, Le Son; Nguyen, Tam T K; Mittal, Deepak; Villani, Rehan; Steptoe, Raymond J; Khosrotehrani, Kiarash; Berzins, Stuart P; Baxter, Alan G; Godfrey, Dale I; Blumenthal, Antje

    2018-01-01

    Natural killer T (NKT) cells are prominent innate-like lymphocytes in the liver with critical roles in immune responses during infection, cancer, and autoimmunity. Interferon gamma (IFN-γ) and IL-4 are key cytokines rapidly produced by NKT cells upon recognition of glycolipid antigens presented by antigen-presenting cells (APCs). It has previously been reported that the transcriptional coactivator β-catenin regulates NKT cell differentiation and functionally biases NKT cell responses toward IL-4, at the expense of IFN-γ production. β-Catenin is not only a central effector of Wnt signaling but also contributes to other signaling networks. It is currently unknown whether Wnt ligands regulate NKT cell functions. We thus investigated how Wnt ligands and β-catenin activity shape liver NKT cell functions in vivo in response to the glycolipid antigen, α-galactosylceramide (α-GalCer) using a mouse model. Pharmacologic targeting of β-catenin activity with ICG001, as well as myeloid-specific genetic ablation of Wntless (Wls) , to specifically target Wnt protein release by APCs, enhanced early IFN-γ responses. By contrast, within several hours of α-GalCer challenge, myeloid-specific Wls deficiency, as well as pharmacologic targeting of Wnt release using the small molecule inhibitor IWP-2 impaired α-GalCer-induced IFN-γ responses, independent of β-catenin activity. These data suggest that myeloid cell-derived Wnt ligands drive early Wnt/β-catenin signaling that curbs IFN-γ responses, but that, subsequently, Wnt ligands sustain IFN-γ expression independent of β-catenin activity. Our analyses in ICG001-treated mice confirmed a role for β-catenin activity in driving early IL-4 responses by liver NKT cells. However, neither pharmacologic nor genetic perturbation of Wnt production affected the IL-4 response, suggesting that IL-4 production by NKT cells in response to α-GalCer is not driven by released Wnt ligands. Collectively, these data reveal complex temporal

  2. Temporal Regulation of Natural Killer T Cell Interferon Gamma Responses by β-Catenin-Dependent and -Independent Wnt Signaling

    Directory of Open Access Journals (Sweden)

    Jessica C. Kling

    2018-03-01

    Full Text Available Natural killer T (NKT cells are prominent innate-like lymphocytes in the liver with critical roles in immune responses during infection, cancer, and autoimmunity. Interferon gamma (IFN-γ and IL-4 are key cytokines rapidly produced by NKT cells upon recognition of glycolipid antigens presented by antigen-presenting cells (APCs. It has previously been reported that the transcriptional coactivator β-catenin regulates NKT cell differentiation and functionally biases NKT cell responses toward IL-4, at the expense of IFN-γ production. β-Catenin is not only a central effector of Wnt signaling but also contributes to other signaling networks. It is currently unknown whether Wnt ligands regulate NKT cell functions. We thus investigated how Wnt ligands and β-catenin activity shape liver NKT cell functions in vivo in response to the glycolipid antigen, α-galactosylceramide (α-GalCer using a mouse model. Pharmacologic targeting of β-catenin activity with ICG001, as well as myeloid-specific genetic ablation of Wntless (Wls, to specifically target Wnt protein release by APCs, enhanced early IFN-γ responses. By contrast, within several hours of α-GalCer challenge, myeloid-specific Wls deficiency, as well as pharmacologic targeting of Wnt release using the small molecule inhibitor IWP-2 impaired α-GalCer-induced IFN-γ responses, independent of β-catenin activity. These data suggest that myeloid cell-derived Wnt ligands drive early Wnt/β-catenin signaling that curbs IFN-γ responses, but that, subsequently, Wnt ligands sustain IFN-γ expression independent of β-catenin activity. Our analyses in ICG001-treated mice confirmed a role for β-catenin activity in driving early IL-4 responses by liver NKT cells. However, neither pharmacologic nor genetic perturbation of Wnt production affected the IL-4 response, suggesting that IL-4 production by NKT cells in response to α-GalCer is not driven by released Wnt ligands. Collectively, these data reveal

  3. 21 CFR 864.7040 - Adenosine triphosphate release assay.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Adenosine triphosphate release assay. 864.7040 Section 864.7040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7040...

  4. IP-10 release assays in the diagnosis of tuberculosis infection

    DEFF Research Database (Denmark)

    Ruhwald, Morten; Aabye, Martine G; Ravn, Pernille

    2012-01-01

    The current state-of-the-art tests for infection with Mycobacterium tuberculosis - the IFN-γ release assays - rely on accurate measurement of the cytokine IFN-γ. Many other potential biomarkers are expressed in concert with IFN-γ, and IP-10 in particular has shown promising results. IP-10...

  5. Proliferative and antiproliferative effects of interferon-gamma and tumor necrosis factor-alpha on cell lines derived from cervical and ovarian malignancies

    International Nuclear Information System (INIS)

    Mutch, D.G.; Massad, L.S.; Kao, M.S.; Collins, J.L.

    1990-01-01

    Four human cell lines derived from cervical carcinomas (ME-180, SiHa, HT-3, and MS751) and three human cell lines derived from ovarian carcinomas (SK-OV-3, Caov-3, and NIH:OVCAR-3) were analyzed in vitro to determine the effect of recombinant interferon-gamma and recombinant human tumor necrosis factor-alpha on cell growth and survival. The effects of interferon-gamma, tumor necrosis factor-alpha, and both interferon-gamma and tumor necrosis factor-alpha on cell growth were measured after 24 and 72 hours of incubation by the incorporation of chromium 51. The results of this analysis showed that all seven cell lines were resistant to the antiproliferative action of tumor necrosis factor-alpha, that the growth of most cell lines was inhibited by interferon-gamma by 72 hours of incubation, and that after 72 hours of incubation all cell lines demonstrated a synergistic antiproliferative response to the combination of interferon-gamma and tumor necrosis factor-alpha. However, the effects of these cytokines on cell growth were found to differ among cell lines and varied with the concentration and the duration of incubation. The growth of one cell line (Caov-3) was stimulated by both tumor necrosis factor-alpha and interferon-gamma. These results suggest that the clinical effects of these cytokines on the growth of gynecologic cancers may be more complex than previously supposed

  6. Adjuvant interferon gamma in patients with drug – resistant pulmonary tuberculosis: a pilot study

    Directory of Open Access Journals (Sweden)

    Carbonell Dalia

    2004-10-01

    Full Text Available Abstract Background Tuberculosis (TB is increasing in the world and drug-resistant (DR disease beckons new treatments. Methods To evaluate the action of interferon (IFN gamma as immunoadjuvant to chemotherapy on pulmonary DR-TB patients, a pilot, open label clinical trial was carried out in the Cuban reference ward for the management of this disease. The eight subjects existing in the country at the moment received, as in-patients, 1 × 106 IU of recombinant human IFN gamma intramuscularly, daily for one month and then three times per week up to 6 months as adjuvant to the indicated chemotherapy, according to their antibiograms and WHO guidelines. Sputum samples collection for direct smear observation and culture as well as routine clinical and thorax radiography assessments were done monthly. Results Sputum smears and cultures became negative for acid-fast-bacilli before three months of treatment in all patients. Lesion size was reduced at the end of 6 months treatment; the lesions disappeared in one case. Clinical improvement was also evident; body mass index increased in general. Interferon gamma was well tolerated. Few adverse events were registered, mostly mild; fever and arthralgias prevailed. Conclusions These data suggest that IFN gamma is useful and well tolerated as adjunctive therapy in patients with DR-TB. Further controlled clinical trials are encouraged.

  7. Increase in neutrophil Fc gamma receptor I expression following interferon gamma treatment in rheumatoid arthritis.

    Science.gov (United States)

    Goulding, N J; Knight, S M; Godolphin, J L; Guyre, P M

    1992-04-01

    The therapeutic potential of interferon gamma (IFN gamma) in a number of disease states is still being explored, but progress is hampered by the lack of a suitable measure of in vivo biological activity. To assess the in vivo biological effects of recombinant human IFN gamma (rhIFN gamma), 14 patients were studied in a randomised, prospective, double blind, placebo controlled trial of this cytokine for the treatment of rheumatoid arthritis. The levels of Fc gamma receptors on peripheral blood neutrophils were measured at baseline and after 21 days of once daily, subcutaneous injections of rhIFN gamma or placebo. An induction of neutrophil Fc gamma receptor type I (Fc gamma RI) was seen in the group of patients receiving recombinant human rhIFN gamma but not in those receiving placebo. No change in the expression of Fc gamma RII or Fc gamma RIII was detected. The amount of induction of Fc gamma RI detected on the neutrophils of patients receiving rhIFN gamma did not correlate with clinical measures of response at either 21 days or at the end of the study (24 weeks). No significant clinical responses were observed in the rhIFN gamma group at these times. These data confirm that the reported in vitro effect of IFN gamma on human neutrophil Fc receptor expression can be reproduced in vivo.

  8. Interferon gamma peptidomimetic targeted to hepatic stellate cells ameliorates acute and chronic liver fibrosis in vivo.

    Science.gov (United States)

    Bansal, Ruchi; Prakash, Jai; De Ruiter, Marieke; Poelstra, Klaas

    2014-04-10

    Hepatic stellate cells play a crucial role in the pathogenesis of hepatic fibrosis. Thus, pharmacological inhibition of pro-fibrotic activities of these cells might lead to an effective therapy for this disease. Among the potent anti-fibrotics, interferon gamma (IFNγ), a proinflammatory cytokine, is highly efficacious but it failed in clinical trials due to the poor efficacy and multiple adverse effects attributed to the ubiquitous IFNγ receptor (IFNγR) expression. To resolve these drawbacks, we chemically synthesized a chimeric molecule containing (a) IFNγ signaling peptide (IFNγ peptidomimetic, mimγ) that retains the agonistic activities of IFNγ but lacks an extracellular receptor recognition sequence for IFNγR; coupled via heterobifunctional PEG linker to (b) bicyclic platelet derived growth factor beta receptor (PDGFβR)-binding peptide (BiPPB) to induce internalization into the stellate cells that express PDGFβR. The synthesized targeted IFNγ peptidomimetic (mimγ-BiPPB) was extensively investigated for its anti-fibrotic and adverse effects in acute and chronic CCl4-induced liver fibrosis models in mice. Treatment with mimγ-BiPPB, after the onset of disease, markedly inhibited both early and established hepatic fibrosis as reflected by a reduced intrahepatic α-SMA, desmin and collagen-I mRNA expression and protein levels. While untargeted mimγ and BiPPB had no effect, and native IFNγ only induced a moderate reduction. Additionally, no off-target effects, e.g. systemic inflammation, were found with mimγ-BiPPB, which were substantially observed in mice treated with native IFNγ. The present study highlights the beneficial effects of a novel BiPPB mediated cell-specific targeting of IFNγ peptidomimetic to the disease-inducing cells and therefore represents a highly potential therapeutic approach to treat fibrotic diseases. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Development of an aptamer beacon for detection of interferon-gamma.

    Science.gov (United States)

    Tuleuova, Nazgul; Jones, Caroline N; Yan, Jun; Ramanculov, Erlan; Yokobayashi, Yohei; Revzin, Alexander

    2010-03-01

    Traditional antibody-based affinity sensing strategies employ multiple reagents and washing steps and are unsuitable for real-time detection of analyte binding. Aptamers, on the other hand, may be designed to monitor binding events directly, in real-time, without the need for secondary labels. The goal of the present study was to design an aptamer beacon for fluorescence resonance energy transfer (FRET)-based detection of interferon-gamma (IFN-gamma)--an important inflammatory cytokine. Variants of DNA aptamer modified with biotin moieties and spacers were immobilized on avidin-coated surfaces and characterized by surface plasmon resonance (SPR). The SPR studies showed that immobilization of aptamer via the 3' end resulted in the best binding IFN-gamma (K(d) = 3.44 nM). This optimal aptamer variant was then used to construct a beacon by hybridizing fluorophore-labeled aptamer with an antisense oligonucleotide strand carrying a quencher. SPR studies revealed that IFN-gamma binding with an aptamer beacon occurred within 15 min of analyte introduction--suggesting dynamic replacement of the quencher-complementary strand by IFN-gamma molecules. To further highlight biosensing applications, aptamer beacon molecules were immobilized inside microfluidic channels and challenged with varying concentration of analyte. Fluorescence microscopy revealed low fluorescence in the absence of analyte and high fluorescence after introduction of IFN-gamma. Importantly, unlike traditional antibody-based immunoassays, the signal was observed directly upon binding of analyte without the need for multiple washing steps. The surface immobilized aptamer beacon had a linear range from 5 to 100 nM and a lower limit of detection of 5 nM IFN-gamma. In conclusion, we designed a FRET-based aptamer beacon for monitoring of an inflammatory cytokine-IFN-gamma. In the future, this biosensing strategy will be employed to monitor dynamics of cytokine production by the immune cells.

  10. Secretion of interferon gamma from human immune cells is altered by exposure to tributyltin and dibutyltin.

    Science.gov (United States)

    Lawrence, Shanieek; Reid, Jacqueline; Whalen, Margaret

    2015-05-01

    Tributyltin (TBT) and dibutyltin (DBT) are widespread environmental contaminants found in food, beverages, and human blood samples. Both of these butyltins (BTs) interfere with the ability of human natural killer (NK) cells to lyse target cells and alter secretion of the pro-inflammatory cytokine tumor necrosis factor alpha (TNFα) from human immune cells in vitro. The capacity of BTs to interfere with secretion of other pro-inflammatory cytokines has not been examined. Interferon gamma (IFNγ) is a modulator of adaptive and innate immune responses, playing an important role in overall immune competence. This study shows that both TBT and DBT alter secretion of IFNγ from human immune cells. Peripheral blood cell preparations that were increasingly reconstituted were used to determine if exposures to either TBT or DBT affected IFNγ secretion and how the makeup of the cell preparation influenced that effect. IFNγ secretion was examined after 24 h, 48 h, and 6 day exposures to TBT (200 - 2.5 nM) and DBT (5 - 0.05 µM) in highly enriched human NK cells, a monocyte-depleted preparation of PBMCs, and monocyte-containing PBMCs. Both BTs altered IFNγ secretion from immune cells at most of the conditions tested (either increasing or decreasing secretion). However, there was significant variability among donors as to the concentrations and time points that showed changes as well as the baseline secretion of IFNγ. The majority of donors showed an increase in IFNγ secretion in response to at least one concentration of TBT or DBT at a minimum of one length of exposure. © 2013 Wiley Periodicals, Inc.

  11. [Adenovirus-mediated canine interferon-gamma expression and its antiviral activity against canine parvovirus].

    Science.gov (United States)

    Zhang, Kao; Jin, Huijun; Zhong, Fei; Li, Xiujin; Neng, Changai; Chen, Huihui; Li, Wenyan; Wen, Jiexia

    2012-11-04

    To construct recombinant adenovirus containing canine interferon-gamma (cIFN-gamma) gene and to investigate its antiviral activity against canine parvovirus in Madin-Darby canine kidney cells (MDCK). [Methods] The cIFN-gamma gene was inserted into adenovirus shuttle plasmid to construct pShuttle3-cIFN-gamma expression vector, from which the cIFN-gamma expression cassette was transferred into the adenovirus genomic plasmid pAdeno-X by specific restriction sites to generate recombinant adenovirus genomic plasmid pAd-cIFN-gamma. The pAd-cIFN-gamma plasmid was linearized by digestion and transfected into human embryonic kidney (HEK) 293T cells to generate the replication-defective cIFN-gamma recombinant adenovirus (Ad-cIFN-gamma). To analyze its anti-canine parvovirus activity, the MDCK cells were pre-infected by Ad-cIFN-gamma recombinant adenovirus, and then infected by canine parvovirus. The antiviral activity of the Ad-cIFN-gamma recombinant adenovirus against parvovirus was analyzed. The recombinant adenovirus containing cIFN-gamma gene was constructed by the ligation method. The recombinant adenovirus could mediates recombinant cIFN-gamma secretory expression in MDCK cells. The Ad-cIFN-gamma recombinant adenovirus could significantly inhibit canine parvovirus replication in MDCK cells pre-infected with the recombinant adenovirus. These results indicate that the Ad-cIFN-gamma recombinant adenovirus has the potent antiviral activity against canine parvovirus. The Ad-cIFN-gamma recombinant adenovirus was successfully constructed by the ligation method and possessed a powerful antiviral activity against canine parvovirus.

  12. Heterogeneity within populations of recombinant Chinese hamster ovary cells expressing human interferon-gamma.

    Science.gov (United States)

    Coppen, S R; Newsam, R; Bull, A T; Baines, A J

    1995-04-20

    The Chinese hamster ovary (CHO) cell line has great commercial importance in the production of recombinant human proteins, especially those for therapeutic use. Much attention has been paid to CHO cell population physiology in order to define factors affecting product fidelity and yield. Such studies have revealed that recombinant proteins, including human interferon-gamma (IFN-gamma), can be heterogeneous both in glycosylation and in proteolytic processing. The type of heterogeneity observed depends on the growth physiology of the cell population, although the relationship between them is complex. In this article we report results of a cytological study of the CHO320 line which expresses recombinant human IFN-gamma. When grown in suspension culture, this cell line exhibited three types of heterogeneity: (1) heterogeneity of the production of IFN-gamma within the cell population, (2) heterogeneity of the number of nuclei and mitotic spindles in dividing cells, and (3) heterogeneity of cellular environment. The last of these arises from cell aggregates which form in suspension culture: Some cells are exposed to the culture medium; others are fully enclosed within the mass with little or no direct access to the medium. Thus, live cells producing IFN-gamma are heterogeneous in their environment, with variable access to O(2) and nutrients. Within the aggregates, it appears that live cells proliferate on a dead cell mass. The layer of live cells can be several cells deep. Specific cell-cell attachments are observed between the living cells in these aggregates. Two proteins, known to be required for the formation of certain types of intercellular junctions, spectrin and vinculin, have been localized to the regions of cell-cell contact. The aggregation of the cells appears to be an active process requiring protein synthesis. (c) 1995 John Wiley & Sons, Inc.

  13. Is TB Testing Associated With Increased Blood Interferon-Gamma Levels?

    Directory of Open Access Journals (Sweden)

    Aideen E. Kennedy

    2017-10-01

    Full Text Available The Republic of Ireland reports a relatively low prevalence of Johne’s disease (JD compared to international counterparts. Postulated reasons for this include a lower average herd size and a grass-based production system. Ireland also engages in high levels of bovine tuberculosis (bTB testing. As interferon-gamma (IFN-γ is believed to play a key role in protecting against JD, it is our hypothesis that administration of purified protein derivative (PPD, as part of the bTB test, is associated with a systemic increase in IFN-γ production, which may potentially limit clinical progression of the disease. We studied 265 cows (202 Friesian and 63 “Non-Friesian,” e.g., JerseyX, Norwegian Red to assess IFN-γ levels and Mycobacterium avium subspecies paratuberculosis (MAP antibody response before and after the bTB test. As part of the compulsory annual bTB test, avian and bovine PPD were administered at two separate cervical sites. To assess IFN-γ production, blood samples were taken before and 72 h after PPD administration. MAP antibody response was assessed before and 10 days post-PPD administration. A significant increase in MAP antibody response was identified post-bTB compared to pre-bTB response (p < 0.001. Additionally, IFN-γ production significantly increased at the post-bTB time point (p < 0.001 compared to the pre-bTB test readings. This may indicate a beneficial effect of bTB testing in controlling JD.

  14. Interferon-gamma inducible protein-10 as a potential biomarker in localized scleroderma

    Science.gov (United States)

    2013-01-01

    Introduction The purpose of this study was to evaluate the presence and levels of interferon-gamma inducible protein-10 (IP-10) in the plasma and skin of pediatric localized scleroderma (LS) patients compared to those of healthy pediatric controls and to determine if IP-10 levels correlate to clinical disease activity measures. Methods The presence of IP-10 in the plasma was analyzed using a Luminex panel in 69 pediatric patients with LS and compared to 71 healthy pediatric controls. Of these patients, five had available skin biopsy specimens with concurrent clinical and serological data during the active disease phase, which were used to analyze the presence and location of IP-10 in the skin by immunohistochemistry (IHC). Results IP-10 levels were significantly elevated in the plasma of LS patients compared to that of healthy controls and correlated to clinical disease activity measures in LS. Immunohistochemistry staining of IP-10 was present in the dermal infiltrate of LS patients and was similar to that found in psoriasis skin specimens, the positive disease control. Conclusions Elevation of IP-10 levels in the plasma compared to those of healthy controls and the presence of IP-10 staining in the affected skin of LS patients indicates that IP-10 is a potential biomarker in LS. Furthermore, significant elevation of IP-10 in LS patients with active versus inactive disease and correlations between IP-10 levels and standardized disease outcome measures of activity in LS strongly suggest that IP-10 may be a biomarker for disease activity in LS. PMID:24499523

  15. Interferon-gamma sensitizes colonic epithelial cell lines to physiological and therapeutic inducers of colonocyte apoptosis.

    LENUS (Irish Health Repository)

    O'Connell, J

    2012-02-03

    Homeostasis in the colonic epithelium is achieved by a continuous cycle of proliferation and apoptosis, in which imbalances are associated with disease. Inflammatory bowel disease (IBD) and colon cancer are associated with either excessive or insufficient apoptosis of colonic epithelial cells, respectively. By using two colonic epithelial cell lines, HT29 and SW620, we investigated how the epithelial cell\\'s sensitivity to apoptosis was regulated by the proinflammatory cytokine interferon-gamma (IFN-gamma). We found that IFN-gamma sensitized HT29 cells, and to a lesser extent SW620, to diverse inducers of apoptosis of physiologic or therapeutic relevance to the colon. These apoptosis inducers included Fas (CD95\\/APO-1) ligand (FasL), short-chain fatty acids, and chemotherapeutic drugs. The extent of IFN-gamma-mediated apoptosis sensitization in these two cell lines correlated well with the degree of IFN-gamma-mediated upregulation of the proapoptotic protease caspase-1. Although IFN-gamma alone effectively sensitized HT29 cells to apoptosis, inclusion of the protein synthesis inhibitor cyclohexamide (CHX) during apoptotic challenge was necessary for maximal sensitization of SW620. The requirement of CHX to sensitize SW620 cells to apoptosis implies a need to inhibit translation of antiapoptotic proteins absent from HT29. In particular, the antiapoptotic protein Bcl-2 was strongly expressed in SW620 cells but absent from HT29. Our results indicate that IFN-gamma increases the sensitivity of colonic epithelial cells to diverse apoptotic stimuli in concert, via upregulation of caspase-1. Our findings implicate caspase-1 and Bcl-2 as important central points of control determining the general sensitivity of colonic epithelial cells to apoptosis.

  16. Interferon-gamma and interleukin-10 profile of children with tuberculosis in North Sumatera, Indonesia

    Science.gov (United States)

    Daulay, R. S.; Daulay, R. M.

    2018-03-01

    Cellular immunity was mediated the host immune response against Mycobacterium tuberculosis, in which cytokine and T-helper (Th) 1 cells play an important role. Interferon-gamma (IFN-γ) is a leading cytokine involved in the immune response of tuberculosis (TB).The primary function of IFN-γ is to activate macrophages in opposition Mycobacterium tuberculosis. Contrast from IFN-γ, interleukin-10 (IL-10) is considered inhibitory cytokine, important to an adequate balance between inflammatory responses. To analyze cytokine profile, particularly IFN-γ and IL-10 of the children with TB in Indonesia, a cross-sectional study was conducted at two general hospitals and seven primary health care located in Medan and Batubara, North Sumatera, Indonesia. Among 51 children with TB disease and 51 healthy children, found that IFN-γ and IL-10 levels were lower in TB patients compared to healthy children. Statistically significant decreased production of the IFN-γ levels (p=0.042) were found in TB patients 9.41 (1.10-28.06) pg/ml contrast to healthy children 6.30 (1.30-89.76) pg/ml. Homologue finding of the IL-10 levels were also found in TB patients 4.93 (0.22-48.01) pg/ml and 4.93 (0.07-81.60) pg/ml in healthy children, but not statistically significant (p=0.784). High levels of IL-10 were not proven to suppress the levels production of IFN-γ in TB patients.

  17. Functional polymorphisms of interferon-gamma affect pneumonia-induced sepsis.

    Directory of Open Access Journals (Sweden)

    Ding Wang

    Full Text Available Sepsis is an inflammatory syndrome caused by infection, and both its incidence and mortality are high. Because interferon-gamma (IFN-γ plays an important role in inflammation, this work assessed IFN-γ single nucleotide polymorphism (SNPs that may be associated with sepsis.A total of 196 patients with pneumonia-induced sepsis and 213 age- and sex-matched healthy volunteers participated in our study from July 2012 to July 2013 in Guangzhou, China. Patient clinical information was collected. Clinical pathology was assessed in subgroups defined based on clinical criteria, APACHE II (acute physiology and chronic health evaluation and SOFA (sepsis-related organ failure assessment scores and discharge rate. Four functional SNPs, -1616T/C (rs2069705, -764G/C (rs2069707, +874A/T (rs2430561 and +3234C/T (rs2069718, were genotyped by Snapshot in both sepsis patients and healthy controls. Pearson's chi-square test or Fisher's exact test were used to analyze the distribution of the SNPs, and the probability values (P values, odds ratios (OR and 95% confidence intervals (CIs were calculated.No mutations in the IFN-γ -764G/C SNP were detected among the participants in our study. The +874A/T and +3234C/T SNPs were in strong linkage disequilibrium (LD (r(2 = 0.894. The -1616 TC+TT, +874 AT+AA genotype and the TAC haplotype were significantly associated with sepsis susceptibility, while the CTT haplotype was associated with protection against sepsis incidence. Genotype of -1616 TT wasn't only protective against severity of sepsis, but also against higher APACHE II and SOFA scores as +874 AA and +3234 CC. The TAC haplotype was was protective against progression to severe sepsis either.Our results suggest that functional IFN-γ SNPs and their haplotypes are associated with pneumonia-induced sepsis.

  18. Growth hormone, interferon-gamma, and leukemia inhibitory factor utilize insulin receptor substrate-2 in intracellular signaling

    DEFF Research Database (Denmark)

    Argetsinger, L S; Norstedt, G; Billestrup, Nils

    1996-01-01

    In this report, we demonstrate that insulin receptor substrate-2 (IRS-2) is tyrosyl-phosphorylated following stimulation of 3T3-F442A fibroblasts with growth hormone (GH), leukemia inhibitory factor and interferon-gamma. In response to GH and leukemia inhibitory factor, IRS-2 is immediately...... for GH is further demonstrated by the finding that GH stimulates association of IRS-2 with the 85-kDa regulatory subunit of phosphatidylinositol 3'-kinase and with the protein-tyrosine phosphatase SHP2. These results are consistent with the possibility that IRS-2 is a downstream signaling partner...

  19. Evaluation of the prognostic value of IFN-gamma release assay and tuberculin skin test in household contacts of infectious tuberculosis cases in Senegal.

    Directory of Open Access Journals (Sweden)

    Christian Lienhardt

    2010-05-01

    Full Text Available Chemoprophylaxis of contacts of infectious tuberculosis (TB cases is recommended for TB control, particularly in endemic countries, but is hampered by the difficulty to diagnose latent TB infection (LTBI, classically assessed through response to the Tuberculin Skin Test (TST. Interferon-gamma release assays (IGRA are proposed new tools to diagnose LTBI, but there are limited data on their ability to predict the development of active TB disease. To address this, we investigated the response to TST and IGRA in household contacts of infectious TB cases in a TB high-burden country and the potential correlation with development of TB.Prospective household contacts study conducted in two health centres in Dakar, Senegal. A total of 2679 household contacts of 206 newly detected smear and/or culture positive index TB cases aged 18 years or greater were identified A TST was performed in each contact and an ESAT6/CFP10 ELISPOT assay performed in a random sample of those. Contacts were followed-up for 24 months. TB was diagnosed in 52 contacts, an incidence rate of 9.27/1000 person-years. In univariable analysis, the presence of positive TST (> or = 10 mm and ELISPOT (>32 SFC/million PBMC responses at baseline were associated with active TB during follow-up: Rate Ratio [RR] = 2.32 (95%CI:1.12-4.84 and RR = 2.09 (95%CI:0.83-5.31, respectively. After adjustment for age, sex and proximity to index case, adjusted RRs were 1.51 (95%CI:0.71-3.19 and 1.98 (95%CI:0.77-5.09, respectively. Restricting analysis to the 40 microbiologically confirmed cases, the adjusted RR for positive ELISPOT was 3.61 (95%CI:1.03-12.65. The median ELISPOT response in contacts who developed TB was 5-fold greater than in those who did not develop TB (p = 0.02.TST and IGRAs are markers of a contact of the immune system with tubercle bacilli. In a TB endemic area, a high ELISPOT response may reflect increased bacterial replication that may subsequently be associated with development of TB

  20. A highly sensitive label-free electrochemical aptasensor for interferon-gamma detection based on graphene controlled assembly and nuclease cleavage-assisted target recycling amplification.

    Science.gov (United States)

    Yan, Genping; Wang, Yonghong; He, Xiaoxiao; Wang, Kemin; Liu, Jinquan; Du, Yudan

    2013-06-15

    We report here a highly sensitive and label-free electrochemical aptasensing technology for detection of interferon-gamma (IFN-γ) based on graphene controlled assembly and enzyme cleavage-assisted target recycling amplification strategy. In this work, in the absence of IFN-γ, the graphene could not be assembled onto the 16-mercaptohexadecanoic acid (MHA) modified gold electrode because the IFN-γ binding aptamer was strongly adsorbed on the graphene due to the strong π-π interaction. Thus the electronic transmission was blocked (eT OFF). However, the presence of target IFN-γ and DNase I led to desorption of aptamer from the graphene surface and further cleavage of the aptamer, thereby releasing the IFN-γ. The released IFN-γ could then re-attack other aptamers on the graphene, resulting in the successive release of the aptamers from the graphene. At the same time, the "naked" graphene could be assembled onto the MHA modified gold electrode with hydrophobic interaction and π-conjunction, mediating the electron transfer between the electrode and the electroactive indicator. Then, measurable electrochemical signals were generated (eT ON), which was related to the concentration of the IFN-γ. By taking advantages of graphene and enzyme cleavage-assisted target recycling amplification, the developed label-free electrochemical aptasensing technology showed a linear response to concentration of IFN-γ range from 0.1 to 0.7 pM. The detection limit of IFN-γ was determined to be 0.065 pM. Moreover, this aptasensor shows good selectivity toward the target in the presence of other relevant proteins. Our strategy thus opens new opportunities for label-free and amplified detection of other kinds of proteins. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. The effect of co-administration of DNA carrying chicken interferon-gamma gene on protection of chickens against infectious bursal disease by DNA-mediated vaccination.

    Science.gov (United States)

    Hsieh, Ming Kun; Wu, Ching Ching; Lin, Tsang Long

    2006-11-17

    The purpose of the present study was to determine whether DNA vaccination by co-administration of DNA coding for chicken interferon-gamma (IFN-gamma) gene and DNA encoding for the VP243 gene of IBDV could enhance immune response and protection efficacy of chickens against challenge by IBDV. Plasmids carrying VP243 gene of IBDV strain variant E (VE) (P/VP243/E) and chicken IFN-gamma gene (P/cIFN-gamma) were constructed, respectively. One-day-old chickens were intramuscularly injected with P/VP243/E, or P/cIFN-gamma, or both once, twice, or three times into the thigh muscle of one leg or the thigh muscles of two separate legs at weekly intervals. Chickens were orally challenged with IBDV strain VE at 3 weeks of age and observed for 10 days. Chickens receiving two plasmids in the same site two times had significantly higher (Pprotection and those receiving two plasmids in the same sites did not have any protection against IBD. The enzyme-linked immunosorbent assay (ELISA) and virus neutralization (VN) titers to IBDV of chickens in the groups with three doses of P/VP243/E were significantly higher (Pprotected by DNA vaccination did not have detectable IBDV antigen in the bursae as determined by immunofluorescent antibody assay (IFA). The results indicated that co-administration of plasmid encoding chicken IFN-gamma gene with plasmid encoding a large segment gene of the IBDV did not enhance immune response and protection against challenge by IBDV.

  2. Marrow stem cell release in the autorepopulation assay

    Energy Technology Data Exchange (ETDEWEB)

    Maloney, M A; Patt, H M [California Univ., San Francisco (USA). Lab. of Radiobiology

    1978-01-01

    The early migration of stem cells from shielded marrow to an irradiated spleen has been re-evaluated, and the findings have been compared with the results of earlier studies. The composite data reveal a constant rate during the first 24 h after irradiation, with a slope of 1.6 cells per h and an intercept of 2.4. The positive intercept is interpreted to signify an immediate brief perturbation of CFU/sub s/ release. The low concentration of CFU/sub s/ in the bloodstream, despite their continuous migration from the shielded marrow, is indicative of a rapid, and probably greatly increased, blood turnover. Despite the constancy of stem cell seeding, it is not yet possible to determine whether the rate of stem cell release is different in shielded marrow than in normal marrow. The resolution of this question requires more precise information about spleen seeding efficiency in the autorepopulation assay and about the normal turnover rate of stem cells in the bloodstream.

  3. Cross-talk between IGF-1 and estrogen receptors attenuates intracellular changes in ventral spinal cord 4.1 motoneuron cells due to interferon-gamma exposure

    Science.gov (United States)

    Park, Sookyoung; Nozaki, Kenkichi; Smith, Joshua A.; Krause, James S.; Banik, Naren L.

    2014-01-01

    Insulin-like growth factor-1 (IGF-1) is a neuroprotective growth factor that promotes neuronal survival by inhibition of apoptosis. In order to examine whether IGF-1 exerts cytoprotective effects against extracellular inflammatory stimulation, ventral spinal cord 4.1 (VSC4.1) motoneuron cells were treated with interferon-gamma (IFN-γ). Our data demonstrated apoptotic changes, increased calpain:calpastatin and Bax:Bcl-2 ratios, and expression of apoptosis related proteases (caspase-3 and −12) in motoneurons rendered by IFN-γ in a dose-dependent manner. Post-treatment with IGF-1 attenuated these changes. In addition, IGF-1 treatment of motoneurons exposed to IFN-γ decreased expression of inflammatory markers (cyclooxygenase-2 and nuclear factor-kappa B:inhibitor of kappa B ratio). Furthermore, IGF-1 attenuated the loss of expression of IGF-1 receptors (IGF-1Rα and IGF-1Rβ) and estrogen receptors (ERα and ERβ) induced by IFN-γ. To determine whether the protective effects of IGF-1 are associated with ERs, ERs antagonist ICI and selective siRNA targeted against ERα and ERβ were used in VSC4.1 motoneurons. Distinctive morphological changes were observed following siRNA knockdown of ERα and ERβ. In particular, apoptotic cell death assessed by TUNEL assay was enhanced in both ERα and ERβ-silenced VSC4.1 motoneurons following IFN-γ and IGF-1 exposure. These results suggest that IGF-1 protects motoneurons from inflammatory insult by a mechanism involving pivotal interactions with ERα and ERβ. PMID:24188094

  4. Cross-talk between IGF-1 and estrogen receptors attenuates intracellular changes in ventral spinal cord 4.1 motoneuron cells because of interferon-gamma exposure.

    Science.gov (United States)

    Park, Sookyoung; Nozaki, Kenkichi; Smith, Joshua A; Krause, James S; Banik, Naren L

    2014-03-01

    Insulin-like growth factor-1 (IGF-1) is a neuroprotective growth factor that promotes neuronal survival by inhibition of apoptosis. To examine whether IGF-1 exerts cytoprotective effects against extracellular inflammatory stimulation, ventral spinal cord 4.1 (VSC4.1) motoneuron cells were treated with interferon-gamma (IFN-γ). Our data demonstrated apoptotic changes, increased calpain:calpastatin and Bax:Bcl-2 ratios, and expression of apoptosis-related proteases (caspase-3 and -12) in motoneurons rendered by IFN-γ in a dose-dependent manner. Post-treatment with IGF-1 attenuated these changes. In addition, IGF-1 treatment of motoneurons exposed to IFN-γ decreased expression of inflammatory markers (cyclooxygenase-2 and nuclear factor-kappa B:inhibitor of kappa B ratio). Furthermore, IGF-1 attenuated the loss of expression of IGF-1 receptors (IGF-1Rα and IGF-1Rβ) and estrogen receptors (ERα and ERβ) induced by IFN-γ. To determine whether the protective effects of IGF-1 are associated with ERs, ERs antagonist ICI and selective siRNA targeted against ERα and ERβ were used in VSC4.1 motoneurons. Distinctive morphological changes were observed following siRNA knockdown of ERα and ERβ. In particular, apoptotic cell death assessed by TUNEL assay was enhanced in both ERα and ERβ-silenced VSC4.1 motoneurons following IFN-γ and IGF-1 exposure. These results suggest that IGF-1 protects motoneurons from inflammatory insult by a mechanism involving pivotal interactions with ERα and ERβ. © 2013 International Society for Neurochemistry.

  5. A synthetic NCAM-derived mimetic peptide, FGL, exerts anti-inflammatory properties via IGF-1 and interferon-gamma modulation

    DEFF Research Database (Denmark)

    Downer, Eric J; Cowley, Thelma R; Cox, Fionnuala

    2009-01-01

    activation and subsequent pro-inflammatory cytokine production. The aim of the current study was to determine if FGL corrects the age-related imbalance in hippocampal levels of insulin-like growth factor-1 (IGF-1) and pro-inflammatory interferon-gamma (IFNgamma), and subsequently attenuates the glial...

  6. 5HT(4) agonists inhibit interferon-gamma-induced MHC class II and B7 costimulatory molecules expression on cultured astrocytes

    NARCIS (Netherlands)

    Zeinstra, Esther M.; Wilczak, Nadine; Wilschut, Jan C.; Glazenburg, Lisa; Chesik, Daniel; Kroese, Frans G. M.; De Keyser, Jacques

    2006-01-01

    A failure of tight control of MHC class II expression on astrocytes may play a role in the development of autoimmune responses in multiple sclerosis. The 5-HT4 serotonin receptor agonists cisapride and prucalopride, at concentrations between 10(-10) M and 10(-8) M, reduced interferon-gamma-induced

  7. Effects of interferon gamma and specific polyclonal antibody on the infection of murine peritoneal macrophages and murine macrophage cell line PMJ2-R with Encephalitozoon cuniculi

    Czech Academy of Sciences Publication Activity Database

    Jelínek, Jiří; Salát, Jiří; Sak, Bohumil; Kopecký, Jan

    2007-01-01

    Roč. 54, č. 3 (2007), s. 172-176 ISSN 0015-5683 R&D Projects: GA ČR GP524/03/D167 Institutional research plan: CEZ:AV0Z60220518 Keywords : microsporidia * Encephalitozoon cuniculi * antibody * macrophage s * interferon gamma (IFN-gamma) Subject RIV: EC - Immunology Impact factor: 1.000, year: 2007

  8. Comparison of histopathology, cultivation of tissues and rectal contents, and interferon-gamma and serum antibody responses for the diagnosis of bovine paratuberculosis

    DEFF Research Database (Denmark)

    Huda, A.; Jensen, H.E.

    2003-01-01

    contents, and (3) examination of repeated blood samples for interferon-gamma (IFN-gamma) and antibody responses. Tissue samples were taken from the small and large intestine and corresponding mesenteric lymph nodes, and from the pharyngeal tonsil and other lymphoid nodes (retropharyngeal, mediastinal...

  9. Gamma Interferon Release Assays for Detection of Mycobacterium tuberculosis Infection

    Science.gov (United States)

    Denkinger, Claudia M.; Kik, Sandra V.; Rangaka, Molebogeng X.; Zwerling, Alice; Oxlade, Olivia; Metcalfe, John Z.; Cattamanchi, Adithya; Dowdy, David W.; Dheda, Keertan; Banaei, Niaz

    2014-01-01

    SUMMARY Identification and treatment of latent tuberculosis infection (LTBI) can substantially reduce the risk of developing active disease. However, there is no diagnostic gold standard for LTBI. Two tests are available for identification of LTBI: the tuberculin skin test (TST) and the gamma interferon (IFN-γ) release assay (IGRA). Evidence suggests that both TST and IGRA are acceptable but imperfect tests. They represent indirect markers of Mycobacterium tuberculosis exposure and indicate a cellular immune response to M. tuberculosis. Neither test can accurately differentiate between LTBI and active TB, distinguish reactivation from reinfection, or resolve the various stages within the spectrum of M. tuberculosis infection. Both TST and IGRA have reduced sensitivity in immunocompromised patients and have low predictive value for progression to active TB. To maximize the positive predictive value of existing tests, LTBI screening should be reserved for those who are at sufficiently high risk of progressing to disease. Such high-risk individuals may be identifiable by using multivariable risk prediction models that incorporate test results with risk factors and using serial testing to resolve underlying phenotypes. In the longer term, basic research is necessary to identify highly predictive biomarkers. PMID:24396134

  10. Production of interferon-gamma by in vivo tumor-sensitized T cells: Association with active antitumor immunity

    International Nuclear Information System (INIS)

    Bursuker, I.; Pearce, M.T.

    1990-01-01

    The state of active immunity to Meth A fibrosarcoma in mice immunized with an admixture of Meth A cells and Propionibacterium acnes is associated with possession by the host of spleen cells capable of producing interferon-gamma (IFN-gamma) upon in vitro restimulation with irradiated tumor cells. The ability of spleen cells from immunized mice to produce IFN-gamma in response to irradiated Meth A cells decays as active antitumor immunity is replaced by a state of immunological memory. The IFN-producing cells are L3T4+Ly2+, cyclophosphamide-sensitive and radiosensitive T cells, as determined by their sensitivity to corresponding monoclonal antibodies and complement. The induction of IFN-gamma production by in vivo tumor-sensitized T cells is tumor specific, in that spleen cells from mice immunized against Meth A fibrosarcoma can produce IFN in response to irradiated Meth A cells but not in response to another syngeneic tumor M109 lung carcinoma

  11. Growth hormone, interferon-gamma, and leukemia inhibitory factor promoted tyrosyl phosphorylation of insulin receptor substrate-1

    DEFF Research Database (Denmark)

    Argetsinger, L S; Hsu, G W; Myers, M G

    1995-01-01

    ), the principle substrate of the insulin receptor. Tyrosyl phosphorylation of IRS-1 is a critical step in insulin signaling and provides binding sites for proteins with the appropriate Src homology 2 domains, including the 85-kDa regulatory subunit of phosphatidylinositol (PI) 3'-kinase. In 3T3-F442A fibroblasts......., Campbell, G. S., Allevato, G., Billestrup, N., Norstedt, G., and Carter-Su, C. (1994) J. Biol. Chem. 269, 21709-21717). When other cytokines that activate JAK2 were tested for the ability to stimulate the tyrosyl phosphorylation of IRS-1, stimulation was detected with interferon-gamma and leukemia...... to JAK2. GH is also shown to stimulate binding of IRS-1 to the 85-kDa regulatory subunit of PI 3'-kinase. The ability of GH to stimulate tyrosyl phosphorylation of IRS-1 and its association with PI 3'-kinase provides a biochemical basis for responses shared by insulin and GH including the well...

  12. Potentiating day-old blood samples for detection of interferon-gamma responses following infection with Mycobacterium avium subsp. paratuberculosis

    DEFF Research Database (Denmark)

    Mikkelsen, Heidi; Nielsen, Søren Saxmose; Jungersen, Gregers

    time interval from blood sampling to culture. The objective of the study was to assess options for use of day-old blood samples for early-stage diagnosis of MAP infections. Bovine interleukin 12 (IL-12) can induce, and IL-10 reduce, IFN-γ production. Therefore, addition of IL-12 and anti-IL-10 could...... result in production of IFN-γ in samples previously exposed to MAP antigens. Whole blood samples were collected from heifers in a Danish dairy herd known to be infected with MAP. The samples were collected on three sample dates, and on each date the blood samples were stimulated with PPDj and recombinant......The interferon gamma (IFN-γ) test measuring specific cell-mediated immune responses in whole blood can be used for diagnosis at an early stage of Mycobacterium avium subsp. paratuberculosis (MAP) infection. A major obstacle for the practical use of IFN-γ testing is the recommended maximum 8 hour...

  13. CMOS image sensor for detection of interferon gamma protein interaction as a point-of-care approach.

    Science.gov (United States)

    Marimuthu, Mohana; Kandasamy, Karthikeyan; Ahn, Chang Geun; Sung, Gun Yong; Kim, Min-Gon; Kim, Sanghyo

    2011-09-01

    Complementary metal oxide semiconductor (CMOS)-based image sensors have received increased attention owing to the possibility of incorporating them into portable diagnostic devices. The present research examined the efficiency and sensitivity of a CMOS image sensor for the detection of antigen-antibody interactions involving interferon gamma protein without the aid of expensive instruments. The highest detection sensitivity of about 1 fg/ml primary antibody was achieved simply by a transmission mechanism. When photons are prevented from hitting the sensor surface, a reduction in digital output occurs in which the number of photons hitting the sensor surface is approximately proportional to the digital number. Nanoscale variation in substrate thickness after protein binding can be detected with high sensitivity by the CMOS image sensor. Therefore, this technique can be easily applied to smartphones or any clinical diagnostic devices for the detection of several biological entities, with high impact on the development of point-of-care applications.

  14. Isotope release cytotoxicity assay applicable to human tumors: the use of 111-indium

    Energy Technology Data Exchange (ETDEWEB)

    Frost, P; Wiltrout, R; Maciorowski, Z; Rose, N R

    1977-01-01

    We have demonstrated that human tumors can be labelled efficiently with the 111indium-oxine chelate. Subsequently, this isotope can be released by cytotoxic lymphoid cells. Both natural and induced cytotoxicity can be demonstrated utilizing this isotope release method. Because of the slow spontaneous release of 111indium and its efficient labelling of human tumor cells, this isotope release assay can be utilized in long-term cytotoxic assays in the study of human tumor immunology.

  15. Co-incubation with IL-18 potentiates antigen-specific IFN-γ response in a whole-blood stimulation assay for measurement of cell-mediated immune responses in pigs experimentally infected with Lawsonia intracellularis

    DEFF Research Database (Denmark)

    Riber, Ulla; Boesen, Henriette Toft; Jakobsen, Jeanne Toft

    2011-01-01

    The whole-blood interferon-gamma (IFN-γ) assay is a quantitative in-vitro assay for a direct read out of Ag-specific cell-mediated immune (CMI) responses to infectious diseases. The IFN-γ assay is robust in severe intracellular infections like Brucella or mycobacteria, but more difficult to evalu......The whole-blood interferon-gamma (IFN-γ) assay is a quantitative in-vitro assay for a direct read out of Ag-specific cell-mediated immune (CMI) responses to infectious diseases. The IFN-γ assay is robust in severe intracellular infections like Brucella or mycobacteria, but more difficult...

  16. Diagnosis of Coxiella burnetii infection: comparison of a whole blood interferon-gamma production assay and a Coxiella ELISPOT

    NARCIS (Netherlands)

    Schoffelen, T.; Limonard, G.J.; Bleeker-Rovers, C.P.; Bouwman, J.J.; Meer, J.W. van der; Nabuurs-Franssen, M.H.; Sprong, T.; Deuren, M. van

    2014-01-01

    Diagnosis of ongoing or past infection with Coxiella burnetii, the causative agent of Q fever, relies heavily on serology: the measurement of C. burnetii-specific antibodies, reflecting the host's humoral immune response. However, cell-mediated immune responses play an important, probably even more

  17. Loci controlling lymphocyte production of interferon gamma after alloantigen stimulation in vitro and their co-localization with genes controlling lymphocyte infiltration of tumors and tumor susceptibility

    Czech Academy of Sciences Publication Activity Database

    Lipoldová, Marie; Havelková, Helena; Badalová, Jana; Vojtíšková, Jarmila; Quan, L.; Krulová, Magdalena; Sohrabi, Yahya; Stassen, A. P. M.; Demant, P.

    2010-01-01

    Roč. 59, č. 2 (2010), s. 203-213 ISSN 0340-7004 R&D Projects: GA MŠk(CZ) LC06009; GA AV ČR IAA500520606; GA ČR GD310/08/H077 Institutional research plan: CEZ:AV0Z50520514 Keywords : Tumor susceptibility * Genetic control of interferon gamma production * Lymphocyte infiltration of tumors Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.293, year: 2010

  18. Effects of single-dose atorvastatin on interleukin-6, interferon gamma, and myocardial no-reflow in a rabbit model of acute myocardial infarction and reperfusion

    International Nuclear Information System (INIS)

    Zhao, X.J.; Liu, X.L.; He, G.X.; Xu, H.P.

    2014-01-01

    The mechanisms of statins relieving the no-reflow phenomenon and the effects of single-dose statins on it are not well known. This study sought to investigate the effects of inflammation on the no-reflow phenomenon in a rabbit model of acute myocardial infarction and reperfusion (AMI/R) and to evaluate the effects of single-dose atorvastatin on inflammation and myocardial no-reflow. Twenty-four New Zealand white male rabbits (5-6 months old) were randomized to three groups of eight: a sham-operated group, an AMI/R group, and an atorvastatin-treated group (10 mg/kg). Animals in the latter two groups were subjected to 4 h of coronary occlusion followed by 2 h of reperfusion. Serum levels of interleukin (IL)-6 were measured by enzyme-linked immunosorbent assay. The expression of interferon gamma (IFN-γ) in normal and infarcted (reflow and no-reflow) myocardial tissue was determined by immunohistochemical methods. The area of no-reflow and necrosis was evaluated pathologically. Levels of serum IL-6 were significantly lower in the atorvastatin group than in the AMI/R group (P<0.01). Expression of IFN-γ in infarcted reflow and no-reflow myocardial tissue was also significantly lower in the atorvastatin group than in the AMI/R group. The mean area of no-reflow [47.01% of ligation area (LA)] was significantly smaller in the atorvastatin group than in the AMI/R group (85.67% of LA; P<0.01). The necrosis area was also significantly smaller in the atorvastatin group (85.94% of LA) than in the AMI/R group (96.56% of LA; P<0.01). In a secondary analysis, rabbits in the atorvastatin and AMI/R groups were divided into two groups based on necrosis area (90% of LA): a small group (<90% of LA) and a large group (>90% of LA). There was no significant difference in the area of no-reflow between the small (61.40% of LA) and large groups (69.87% of LA; P>0.05). Single-dose atorvastatin protected against inflammation and myocardial no-reflow and reduced infarct size during AMI/R in

  19. Effects of single-dose atorvastatin on interleukin-6, interferon gamma, and myocardial no-reflow in a rabbit model of acute myocardial infarction and reperfusion

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, X. J. [Affiliated Hospital of Binzhou Medical University, Department of Cardiology, Binzhou, China, Department of Cardiology, Affiliated Hospital of Binzhou Medical University, Binzhou (China); Liu, X. L. [Qilu Hospital, Shandong University, Department of Cardiology, Jinan, China, Department of Cardiology, Qilu Hospital, Shandong University, Jinan (China); He, G. X. [Third Military Medical University, Southwest Hospital, Department of Cardiology, Chongqing, China, Department of Cardiology, Southwest Hospital, Third Military Medical University, Chongqing (China); Xu, H. P. [Affiliated Hospital of Binzhou Medical University, Department of Cardiology, Binzhou, China, Department of Cardiology, Affiliated Hospital of Binzhou Medical University, Binzhou (China)

    2014-03-03

    The mechanisms of statins relieving the no-reflow phenomenon and the effects of single-dose statins on it are not well known. This study sought to investigate the effects of inflammation on the no-reflow phenomenon in a rabbit model of acute myocardial infarction and reperfusion (AMI/R) and to evaluate the effects of single-dose atorvastatin on inflammation and myocardial no-reflow. Twenty-four New Zealand white male rabbits (5-6 months old) were randomized to three groups of eight: a sham-operated group, an AMI/R group, and an atorvastatin-treated group (10 mg/kg). Animals in the latter two groups were subjected to 4 h of coronary occlusion followed by 2 h of reperfusion. Serum levels of interleukin (IL)-6 were measured by enzyme-linked immunosorbent assay. The expression of interferon gamma (IFN-γ) in normal and infarcted (reflow and no-reflow) myocardial tissue was determined by immunohistochemical methods. The area of no-reflow and necrosis was evaluated pathologically. Levels of serum IL-6 were significantly lower in the atorvastatin group than in the AMI/R group (P<0.01). Expression of IFN-γ in infarcted reflow and no-reflow myocardial tissue was also significantly lower in the atorvastatin group than in the AMI/R group. The mean area of no-reflow [47.01% of ligation area (LA)] was significantly smaller in the atorvastatin group than in the AMI/R group (85.67% of LA; P<0.01). The necrosis area was also significantly smaller in the atorvastatin group (85.94% of LA) than in the AMI/R group (96.56% of LA; P<0.01). In a secondary analysis, rabbits in the atorvastatin and AMI/R groups were divided into two groups based on necrosis area (90% of LA): a small group (<90% of LA) and a large group (>90% of LA). There was no significant difference in the area of no-reflow between the small (61.40% of LA) and large groups (69.87% of LA; P>0.05). Single-dose atorvastatin protected against inflammation and myocardial no-reflow and reduced infarct size during AMI/R in

  20. Gut Microbiota in Type 2 Diabetes Individuals and Correlation with Monocyte Chemoattractant Protein1 and Interferon Gamma from Patients Attending a Tertiary Care Centre in Chennai, India

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    Premalatha Pushpanathan

    2016-01-01

    Full Text Available Background: Type 2 diabetes mellitus (T2DM and obesity are associated with changes in gut microbiota and characterized by chronic low-grade inflammation. Monocyte chemoattractant protein-1 (MCP-1 and interferon gamma (IFNγ are proinflammatory cytokines which play an important role in the development of T2DM. We undertook this study to analyze the gut microbiota of T2DM and nondiabetic subjects and to determine the profile of MCP 1 and IFNγ in the same subjects attending a tertiary care center in Chennai, Tamil Nadu, India. Methods: The study included 30 subjects with clinical details. Stool and blood samples were collected from all the subjects. DNA was extracted from fecal samples and polymerase chain reaction was done using fusion primers. Metagenomic analysis was performed using ion torrent sequencing. The reads obtained were in FASTA format and reported as operational taxonomic units. Human MCP 1 and IFNγ enzyme linked immunosorbent assay (ELISA were performed for 23 serum samples. Results: The study consisted of 30 subjects; 17 were T2DM and 13 were nondiabetics. The gut microbiota among T2DM consisted predominantly of Gram negative bacteria; Escherichia and Prevotella, when compared with the nondiabetic group with predominantly Gram positive organisms suchas Faecalibacterium, Eubacterium, and Bifidobacterium. The mean MCP-1 values in the diabetic group were 232.8 pg/ml and in the nondiabetic group 170.84 pg/ml. IFNγ (mean 385.5 pg/ml was raised in glycated hemoglobin (HbA1c group of 6.5–7.5% which was statistically significant. Association of Escherichia with T2DM and association of Bifidobacteria in the nondiabetics were also statistically significant. Conclusion: Escherichia counts were elevated in T2DM with HbA1c of 6.5–8.5% which was statistically significant suggesting that lipopolysaccharides present in the cell wall of Gram-negative bacteria may be responsible for low-grade inflammation as evidenced by elevated MCP-1 and

  1. Serum profile of cytokines interferon gamma and interleukin-10 in ewes subjected to artificial insemination by cervical retraction.

    Science.gov (United States)

    Alvares, C T G; Cruz, J F; Romano, C C; Brandão, F Z

    2016-04-15

    This study evaluated the influence of artificial insemination (AI) by cervical retraction (CRI) on serum levels of interferon gamma (IFNγ) and interleukin-10 (IL-10) in ewes. Synchronized pluriparous Santa Inês ewes were subjected to natural mating (NM, n = 8) and AI, which was performed for a fixed time (55 ± 1 hour) by CRI (n = 8) or laparoscopy (n = 8). Ewes were classified as pregnant, with return to estrus (RE) or with embryonic loss (EL). Blood samples were collected on Day 0, Day 3, Day 5, Day 12, and Day 17 (Day 0 = AI/NM) for progesterone dosage and cytokines were quantified from Day 0 to Day 12. Progesterone levels were constant, except for a decrease in ewes with RE at Day 17 (P ewes with EL had lower serum levels of IFNγ and IL-10 than pregnant ewes and ewes with RE, regardless of the reproductive method used, with averages of 769.1, 714.9, and 555.7 pg/mL for IFNγ and 713.8, 699.3, and 578.7 pg/mL for IL-10 in pregnant ewes, ewes with RE and EL, respectively (P ewes does not alter the profile of serum cytokines IFNγ and IL-10 and does not induce an inflammatory reaction that can compromise pregnancy. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Interferon-gamma improves impaired dentinogenic and immunosuppressive functions of irreversible pulpitis-derived human dental pulp stem cells

    Science.gov (United States)

    Sonoda, Soichiro; Yamaza, Haruyoshi; Ma, Lan; Tanaka, Yosuke; Tomoda, Erika; Aijima, Reona; Nonaka, Kazuaki; Kukita, Toshio; Shi, Songtao; Nishimura, Fusanori; Yamaza, Takayoshi

    2016-01-01

    Clinically, irreversible pulpitis is treated by the complete removal of pulp tissue followed by replacement with artificial materials. There is considered to be a high potential for autologous transplantation of human dental pulp stem cells (DPSCs) in endodontic treatment. The usefulness of DPSCs isolated from healthy teeth is limited. However, DPSCs isolated from diseased teeth with irreversible pulpitis (IP-DPSCs) are considered to be suitable for dentin/pulp regeneration. In this study, we examined the stem cell potency of IP-DPSCs. In comparison with healthy DPSCs, IP-DPSCs expressed lower colony-forming capacity, population-doubling rate, cell proliferation, multipotency, in vivo dentin regeneration, and immunosuppressive activity, suggesting that intact IP-DPSCs may be inadequate for dentin/pulp regeneration. Therefore, we attempted to improve the impaired in vivo dentin regeneration and in vitro immunosuppressive functions of IP-DPSCs to enable dentin/pulp regeneration. Interferon gamma (IFN-γ) treatment enhanced in vivo dentin regeneration and in vitro T cell suppression of IP-DPSCs, whereas treatment with tumor necrosis factor alpha did not. Therefore, these findings suggest that IFN-γ may be a feasible modulator to improve the functions of impaired IP-DPSCs, suggesting that autologous transplantation of IFN-γ-accelerated IP-DPSCs might be a promising new therapeutic strategy for dentin/pulp tissue engineering in future endodontic treatment. PMID:26775677

  3. Innate invariant NKT cells recognize Mycobacterium tuberculosis-infected macrophages, produce interferon-gamma, and kill intracellular bacteria.

    Directory of Open Access Journals (Sweden)

    Isabel Sada-Ovalle

    2008-12-01

    Full Text Available Cellular immunity to Mycobacterium tuberculosis (Mtb requires a coordinated response between the innate and adaptive arms of the immune system, resulting in a type 1 cytokine response, which is associated with control of infection. The contribution of innate lymphocytes to immunity against Mtb remains controversial. We established an in vitro system to study this question. Interferon-gamma is produced when splenocytes from uninfected mice are cultured with Mtb-infected macrophages, and, under these conditions, bacterial replication is suppressed. This innate control of bacterial replication is dependent on CD1d-restricted invariant NKT (iNKT cells, and their activation requires CD1d expression by infected macrophages as well as IL-12 and IL-18. We show that iNKT cells, even in limiting quantities, are sufficient to restrict Mtb replication. To determine whether iNKT cells contribute to host defense against tuberculosis in vivo, we adoptively transferred iNKT cells into mice. Primary splenic iNKT cells obtained from uninfected mice significantly reduce the bacterial burden in the lungs of mice infected with virulent Mtb by the aerosol route. Thus, iNKT cells have a direct bactericidal effect, even in the absence of synthetic ligands such as alpha-galactosylceramide. Our finding that iNKT cells protect mice against aerosol Mtb infection is the first evidence that CD1d-restricted NKT cells mediate protection against Mtb in vivo.

  4. The Tat protein of human immunodeficiency virus-1 enhances hepatitis C virus replication through interferon gamma-inducible protein-10

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    Qu Jing

    2012-04-01

    Full Text Available Abstract Background Co-infection with human immunodeficiency virus-1 (HIV-1 and hepatitis C virus (HCV is associated with faster progression of liver disease and an increase in HCV persistence. However, the mechanism by which HIV-1 accelerates the progression of HCV liver disease remains unknown. Results HIV-1/HCV co-infection is associated with increased expression of interferon gamma-induced protein-10 (IP-10 mRNA in peripheral blood mononuclear cells (PBMCs. HCV RNA levels were higher in PBMCs of patients with HIV-1/HCV co-infection than in patients with HCV mono-infection. HIV-1 Tat and IP-10 activated HCV replication in a time-dependent manner, and HIV-1 Tat induced IP-10 production. In addition, the effect of HIV-1 Tat on HCV replication was blocked by anti-IP-10 monoclonal antibody, demonstrating that the effect of HIV-1 Tat on HCV replication depends on IP-10. Taken together, these results suggest that HIV-1 Tat protein activates HCV replication by upregulating IP-10 production. Conclusions HIV-1/HCV co-infection is associated with increased expression of IP-10 mRNA and replication of HCV RNA. Furthermore, both HIV-1 Tat and IP-10 activate HCV replication. HIV-1 Tat activates HCV replication by upregulating IP-10 production. These results expand our understanding of HIV-1 in HCV replication and the mechanism involved in the regulation of HCV replication mediated by HIV-1 during co-infection.

  5. Interferon-gamma improves impaired dentinogenic and immunosuppressive functions of irreversible pulpitis-derived human dental pulp stem cells.

    Science.gov (United States)

    Sonoda, Soichiro; Yamaza, Haruyoshi; Ma, Lan; Tanaka, Yosuke; Tomoda, Erika; Aijima, Reona; Nonaka, Kazuaki; Kukita, Toshio; Shi, Songtao; Nishimura, Fusanori; Yamaza, Takayoshi

    2016-01-18

    Clinically, irreversible pulpitis is treated by the complete removal of pulp tissue followed by replacement with artificial materials. There is considered to be a high potential for autologous transplantation of human dental pulp stem cells (DPSCs) in endodontic treatment. The usefulness of DPSCs isolated from healthy teeth is limited. However, DPSCs isolated from diseased teeth with irreversible pulpitis (IP-DPSCs) are considered to be suitable for dentin/pulp regeneration. In this study, we examined the stem cell potency of IP-DPSCs. In comparison with healthy DPSCs, IP-DPSCs expressed lower colony-forming capacity, population-doubling rate, cell proliferation, multipotency, in vivo dentin regeneration, and immunosuppressive activity, suggesting that intact IP-DPSCs may be inadequate for dentin/pulp regeneration. Therefore, we attempted to improve the impaired in vivo dentin regeneration and in vitro immunosuppressive functions of IP-DPSCs to enable dentin/pulp regeneration. Interferon gamma (IFN-γ) treatment enhanced in vivo dentin regeneration and in vitro T cell suppression of IP-DPSCs, whereas treatment with tumor necrosis factor alpha did not. Therefore, these findings suggest that IFN-γ may be a feasible modulator to improve the functions of impaired IP-DPSCs, suggesting that autologous transplantation of IFN-γ-accelerated IP-DPSCs might be a promising new therapeutic strategy for dentin/pulp tissue engineering in future endodontic treatment.

  6. Assessment of safety and interferon gamma responses of Mycobacterium bovis BCG vaccine in goat kids and milking goats.

    Science.gov (United States)

    Pérez de Val, Bernat; Vidal, Enric; López-Soria, Sergio; Marco, Alberto; Cervera, Zoraida; Martín, Maite; Mercader, Irene; Singh, Mahavir; Raeber, Alex; Domingo, Mariano

    2016-02-10

    Vaccination of domestic animals has emerged as an alternative long-term strategy for the control of tuberculosis (TB). A trial under field conditions was conducted in a TB-free goat herd to assess the safety of the Mycobacterium bovis BCG vaccine. Eleven kids and 10 milking goats were vaccinated with BCG. Bacterial shedding and interferon gamma (IFN-γ) responses were monitored throughout the study. Comprehensive pathological examination and mycobacterial culture of target tissues were performed. BCG vaccine strain was only isolated from the draining lymph node of the injection site of a kid euthanized at week 8 post-vaccination. The remaining animals were euthanized at week 24. Six out of 20 showed small granulomas at the injection site. BCG shedding was not detected in either faeces or in milk throughout the study. All vaccinated kids showed BCG-induced IFN-γ responses at week 8 post-vaccination. BCG vaccination of goats showed no lack of biological safety for the animals, environment and public health, and local adverse reactions were negligible. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Use of a recombinant vaccinia virus expressing interferon gamma for post-exposure protection against vaccinia and ectromelia viruses.

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    Susan A Holechek

    Full Text Available Post-exposure vaccination with vaccinia virus (VACV has been suggested to be effective in minimizing death if administered within four days of smallpox exposure. While there is anecdotal evidence for efficacy of post-exposure vaccination this has not been definitively studied in humans. In this study, we analyzed post-exposure prophylaxis using several attenuated recombinant VACV in a mouse model. A recombinant VACV expressing murine interferon gamma (IFN-γ was most effective for post-exposure protection of mice infected with VACV and ectromelia virus (ECTV. Untreated animals infected with VACV exhibited severe weight loss and morbidity leading to 100% mortality by 8 to 10 days post-infection. Animals treated one day post-infection had milder symptoms, decreased weight loss and morbidity, and 100% survival. Treatment on days 2 or 3 post-infection resulted in 40% and 20% survival, respectively. Similar results were seen in ECTV-infected mice. Despite the differences in survival rates in the VACV model, the viral load was similar in both treated and untreated mice while treated mice displayed a high level of IFN-γ in the serum. These results suggest that protection provided by IFN-γ expressed by VACV may be mediated by its immunoregulatory activities rather than its antiviral effects. These results highlight the importance of IFN-γ as a modulator of the immune response for post-exposure prophylaxis and could be used potentially as another post-exposure prophylaxis tool to prevent morbidity following infection with smallpox and other orthopoxviruses.

  8. Interferon-gamma increased epithelial barrier function via upregulating claudin-7 expression in human submandibular gland duct epithelium.

    Science.gov (United States)

    Abe, Ayumi; Takano, Kenichi; Kojima, Takashi; Nomura, Kazuaki; Kakuki, Takuya; Kaneko, Yakuto; Yamamoto, Motohisa; Takahashi, Hiroki; Himi, Tetsuo

    2016-06-01

    Tight junctions (TJs) are necessary for salivary gland function and may serve as indicators of salivary gland epithelial dysfunction. IgG4-related disease (IgG4-RD) is a newly recognized fibro-inflammatory condition which disrupts the TJ associated epithelial barrier. The salivary glands are one of the most frequently involved organs in IgG4-RD, however, changes of the TJ associated epithelial barrier in salivary gland duct epithelium is poorly understood. Here, we investigated the regulation and function of TJs in human submandibular gland ductal epithelial cells (HSDECs) in normal and IgG4-RD. We examined submandibular gland (SMG) tissue from eight control individuals and 22 patients with IgG4-RD and established an HSDEC culture system. Immunohistochemistry, immunocytochemistry, western blotting, and measurement of transepithelial electrical resistance (TER) were performed. Claudin-4, claudin-7, occludin, and JAM-A were expressed at the apical side of the duct epithelium in submandibular gland (SMG) tissue and at the cell borders in HSDECs of normal and IgG4-RD. The expression and distribution of TJs in SMG tissue were not different in control individuals and patients with IgG4-RD in vivo and in vitro. Although interferon-gamma (IFNγ) generally disrupts the integrity and function of TJs, as manifested by decreased epithelial barrier function, IFNγ markedly increased the epithelial barrier function of HSDECs via upregulation of claudin-7 expression in HSDECs from patients with IgG4-RD. This is the first report showing an IFNγ-dependent increase in epithelial barrier function in the salivary gland duct epithelium. Our results provide insights into the functional significance of TJs in salivary gland duct epithelium in physiological and pathological conditions, including IgG4-RD.

  9. Secretion of Interferon gamma (IFNγ) from Human Immune Cells is Altered by Exposure to Tributyltin (TBT) and Dibutyltin (DBT)

    Science.gov (United States)

    Lawrence, Shanieek; Reid, Jacqueline; Whalen, Margaret

    2013-01-01

    Tributyltin (TBT) and dibutyltin (DBT) are widespread environmental contaminants found in food, beverages, and human blood samples. Both of these butyltins (BTs) interfere with the ability of human natural killer (NK) cells to lyse target cells and also alter secretion of the pro-inflammatory cytokine tumor necrosis factor alpha (TNFα) from human immune cells in vitro. The capacity of BTs to interfere with secretion of other pro-inflammatory cytokines has not been examined. Interferon gamma (IFNγ) is a modulator of adaptive and innate immune responses, playing an important role in overall immune competence. This study shows that both TBT and DBT alter secretion of IFNγ from human immune cells. Peripheral blood cell preparations that were increasingly reconstituted were used to determine if exposures to either TBT or DBT affected IFNγ secretion and how the makeup of the cell preparation influenced that effect. IFNγ secretion was examined after 24 h, 48 h and 6 day exposures to TBT (200- 2.5 nM) and DBT (5- 0.05 μM) in highly enriched human NK cells, a monocyte-depleted preparation of PBMCs, and monocyte-containing PBMCs. Both BTs altered IFNγ secretion from NK cells at most of the conditions tested (either increasing or decreasing secretion). However, there was significant variability among donors as to the concentrations and time points that showed changes as well as the baseline secretion of IFNγ. The majority of donors showed an increase in IFNγ secretion in response to at least one concentration of TBT or DBT at a minimum of one length of exposure. PMID:24357260

  10. Severe respiratory syncytial virus bronchiolitis in infants is associated with reduced airway interferon gamma and substance P.

    Directory of Open Access Journals (Sweden)

    Malcolm G Semple

    2007-10-01

    Full Text Available Severe human respiratory syncytial virus (hRSV bronchiolitis in previously well infants may be due to differences in the innate immune response to hRSV infection.to determine if factors mediating proposed mechanisms for severe bronchiolitis differ with severity of disease.197 infants admitted to hospital with hRSV bronchiolitis were recruited and grouped according to no oxygen requirement (n = 27, oxygen dependence (n = 114 or mechanical ventilation (n = 56. We collected clinical data, nasopharyngeal aspirate (NPA and if ventilated bronchoalveolar lavage (BAL. Interferon-gamma (IFN-gamma, substance P (SP, interleukin 9 (IL-9, urea and hRSV load, were measured in cell free supernatant from NPA and BAL. Multivariate analysis compared independent effects of clinical, virological and immunological variables upon disease severity. IFN-gamma and SP concentrations were lower in NPA from infants who required oxygen or mechanical ventilation. Viral load and IL-9 concentrations were high but did not vary with severity of disease. Independent predictors of severe disease (in diminishing size of effect were low weight on admission, low gestation at birth, low NPA IFN-gamma and NPA SP. Nasal airway sampling appears to be a useful surrogate for distal airway sampling since concentrations of IFN-gamma, SP, IL-9 and viral load in NPA correlate with the same in BAL.Our data support two proposed mechanisms for severe hRSV disease; reduced local IFN-gamma response and SP mediated inflammation. We found large amounts of hRSV and IL-9 in airways secretions from the upper and lower respiratory tract but could not associate these with disease severity.

  11. Study on Anti-Hepatitis B Surface Antibody Titer and Specific Interferon Gamma Response Among Dentists

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    Manoochehr Makvandi

    2017-01-01

    Full Text Available Background Hepatitis B virus (HBV is a major problem for healthcare workers worldwide, and among them, dentists are at risk of acquiring HBV infection. The prevalence of HBV infection has been reported among the dentists in different regions of the world. Since none of the available drugs can clear HBV infection, the presence of effective immunity against HBV infection is important to prevent HBV infection. Objectives This study aimed at determining HBs antibody and specific HBV gamma interferon among the dentists, who received hepatitis B vaccine. Methods The blood samples were collected from 40 dentists, including 7 endodontics, 2 oral and maxillofacial radiologist, 4 periodontics, 11 oral and maxillofacial surgeons, 6 implantologists, 3 orthodontics, 1 oral and maxillofacial pathologist, 2 esthetic and restorative dentists, and 4 doctors of dental surgery (DDS at from dental college of Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran during December, 2013. Overall, 31 (77.5% dentists had already received 3 doses of recombinant hepatitis B vaccine, and 9 (22.5% had received only two doses of the vaccine. Their sera were tested for HBsAb and anti-HBc-IgG by the Enzyme Linked Immunosorbent Assay (ELISA test. The lymphocyte of individuals was separated from their blood sample by Ficoll-Hypaque, cells were washed with phosphate buffered saline (PBS by centrifugation, and finally the pellet cells was resuspended in RPMI-1640 media. Separated cells were exposed to 2.5 µg of purified recombinant HBs antigen, and supernatants were collected after 72 hours and tested for detection of specific interferon γ level by ELISA test. Results Overall, 97.5% of dentists showed positive HBs antibody test results while 36 showed (90% positive test results for specific interferon γ against hepatitis B virus infection. Conclusions High coverage of 97.5% immune response against hepatitis B infection was found, indicating high efficacy of recombinant

  12. Analysis of chromium-51 release assay data using personal computer spreadsheet software

    International Nuclear Information System (INIS)

    Lefor, A.T.; Steinberg, S.M.; Wiebke, E.A.

    1988-01-01

    The Chromium-51 release assay is a widely used technique to assess the lysis of labeled target cells in vitro. We have developed a simple technique to analyze data from Chromium-51 release assays using the widely available LOTUS 1-2-3 spreadsheet software. This package calculates percentage specific cytotoxicity and lytic units by linear regression. It uses all data points to compute the linear regression and can determine if there is a statistically significant difference between two lysis curves. The system is simple to use and easily modified, since its implementation requires neither knowledge of computer programming nor custom designed software. This package can help save considerable time when analyzing data from Chromium-51 release assays

  13. In vitro assay for ACTH-releasing activity using ACTH radioimmunoassay. ACTH releasing activities by various drugs

    Energy Technology Data Exchange (ETDEWEB)

    Hashimoto, K; Takahara, J; Hosogi, H; Ofuji, N; Yasuhara, T [Okayama Univ. (Japan). School of Medicine

    1976-02-01

    This report deals with an in vitro assay of ACTH releasing activity utilizing pituitary incubation combined with ACTH radioimmunoassay. Half of a rat pituitary was preincubated in 2ml Krebs Ringer bicarbonate buffer containing 0.2% glucose and 0.25% BSA (KRBG-BSA) for 1.5 hr (45 min x 2). The medium was replaced by 1 ml KRBG-BSA and incubated for 30 min. Then the medium was again replaced by 1 ml KRBG-BSA or KRBG-BSA containing test materials and incubated for another 30 min. The amount of ACTH assayed by radioimmunoassay in the 2nd 30 min incubation was compared with that in the 1st 30 min incubation, and the result was expressed as a percentage. In the ACTH radioimmunoassay, anti-ACTH serum was diluted to 1:1,500-3,000. The /sup 125/I-..cap alpha../sup 1 -24/ACTH-antibody system was not affected by lysine-vasopressin (LVP), arginine-vasopressin (AVP), rat's pituitary LH, GH or prolactin. Human /sup 1 -39/ACTH was used as the ACTH standard. The dilution curve of the incubation medium was parallel to the standard curve. Reproducibility of immunoassayable ACTH within-assay was 174 +- 5.0 pg/tube (CV=2.9%). A log dose-relationship was observed between the amounts of stalk median eminence extracts (SME; NIAMDD) added to the incubation medium and its ACTH releasing activities. The sensitivity of this assay method was at least 0.1 SME or 10 mU of LVP and AVP. Using this method, it was found that LVP, AVP, norepinephrine (100 ng/ml--200 ng/ml) and 5-hydroxytryptophane (1 ..mu..g/ml) had ACTH releasing activities, but LH-RH, TRH, glucagon, dopamine, phentolamine, propranolol, haloperidol, prostaglandin E/sub 1/ and indomethacin did not affect the release of ACTH.

  14. Thermostability of IFN-γ and IP-10 release assays for latent infection with Mycobacterium tuberculosis

    DEFF Research Database (Denmark)

    Blauenfeldt, Thomas; Wagner, Dirk; Aabye, Martine Grosos

    2016-01-01

    accuracy of IP-10 release assay and IGRAs. RESULTS: We included 65 patients with confirmed pulmonary tuberculosis and 160 healthy controls from 6 European centres collaborating in the TBnet. In patients, IP-10 responses increased 1.07 (IQR 0.90-1.36) fold and IFN-γ responses decreased 0.88 (IQR 0......INTRODUCTION: Interferon-γ (IFN-γ) inducible protein 10kD (IP-10) and IFN-γ release assays (IGRAs) are immunodiagnostic tests aiming to identify the presence of specific cellular immune responses, interpreted as markers for latent infection with Mycobacterium tuberculosis. Incubation at higher...

  15. A short 2 week dose titration regimen reduces the severity of flu-like symptoms with initial interferon gamma-1b treatment.

    Science.gov (United States)

    Devane, John G; Martin, Mary L; Matson, Mark A

    2014-06-01

    Flu-like symptoms (FLS) are commonly experienced by patients receiving interferon gamma-1b which may cause discontinuation or disruption of dosing during initial therapy or on re-initiation following a break in therapy. In contrast to Type I interferons, the impact of dose-titration on FLS has not been reported and is not a practice described or included in the approved prescribing information for interferon gamma-1b.The objective of this study was to assess the effect of a 2 week titration regimen on the severity of FLS during the initial 3 weeks of therapy with three times weekly subcutaneous injections of interferon gamma-1b. Healthy men and women were randomized into a double-blind, two-period, crossover study. Each study period was 3 weeks in duration and there was a minimum 15 day washout between treatment periods. Two treatment regimens were compared: No Titration dosing (full 50 mcg/m(2) subcutaneously [s.c.] three times weekly for 3 weeks) and Titration (15 mcg/m(2) s.c. three times weekly during week 1, 30 mcg/m(2) s.c. three times weekly during week 2 followed by the full dose of 50 mcg/m(2) s.c. three times weekly during week 3). Subjects remained in the clinic for at least 12 hours following each injection. FLS was based on a composite score for fever, chills, tiredness and muscle aches assessed at baseline and 4, 8 and 12 hours following each injection. Acetaminophen was allowed at the discretion of the PI. The primary endpoint was the change from baseline in FLS severity at 8 hours averaged over the 3 weeks of treatment. Additional endpoints included FLS at 4 and 12 hours, individual flu-like symptoms, rates of discontinuation, incidence of FLS and acetaminophen use. NCT 01929382. Of the 40 subjects randomized, there were 15 (37.5%) discontinuations. Titration resulted in a significant reduction in FLS severity at 8 hours (p = 0.023) averaged over the 3 week treatment period. The difference in 3 week FLS severity reflects differences

  16. Use of the local lymph node assay in assessment of immune function.

    Science.gov (United States)

    van den Berg, Femke A; Baken, Kirsten A; Vermeulen, Jolanda P; Gremmer, Eric R; van Steeg, Harry; van Loveren, Henk

    2005-07-01

    The murine local lymph node assay (LLNA) was originally developed as a predictive test method for the identification of chemicals with sensitizing potential. In this study we demonstrated that an adapted LLNA can also be used as an immune function assay by studying the effects of orally administered immunomodulating compounds on the T-cell-dependent immune response induced by the contact sensitizer 2,4-dinitrochlorobenzene (DNCB). C57Bl/6 mice were treated with the immunotoxic compounds cyclosporin A (CsA), bis(tri-n-butyltin)oxide (TBTO) or benzo[a]pyrene, (B[a]P). Subsequently, cell proliferation and interferon-gamma (IFN-gamma) and interleukin (IL)-4 release were determined in the auricular lymph nodes (LNs) after DNCB application on both ears. Immunosuppression induced by CsA, TBTO and B[a]P was clearly detectable in this application of the LLNA. Cytokine release measurements proved valuable to confirm the results of the cell proliferation assay and to obtain an indication of the effect on Th1/Th2 balance. We believe to have demonstrated the applicability of an adapted LLNA as an immune function assay in the mouse.

  17. A radiolabel-release microwell assay for proteolytic enzymes present in cell culture media

    International Nuclear Information System (INIS)

    Rucklidge, G.J.; Milne, G.

    1990-01-01

    A modified method for the measurement of proteolytic enzyme activity in cell culture-conditioned media has been developed. Using the release of 3H-labeled peptides from 3H-labeled gelatin the method is performed in microwell plates. The substrate is insolubilized and attached to the wells by glutaraldehyde treatment, thus eliminating the need for a precipitation step at the end of the assay. The assay is sensitive, reproducible, and convenient for small sample volumes. The effect of different protease inhibitors on activity can be assessed rapidly allowing an early characterization of the enzyme. It can also be adapted to microplate spectrophotometric analysis by staining residual substrate with Coomassie blue

  18. Dual specific oral tolerance induction using interferon gamma for IgE-mediated anaphylactic food allergy and the dissociation of local skin allergy and systemic oral allergy: tolerance or desensitization?

    Science.gov (United States)

    Noh, G; Jang, E H

    2014-01-01

    Specific oral tolerance induction (SOTI) for IgE-mediated food allergy (IFA) can be successfully achieved using interfero gamma (classic SOTI). In this study, a tolerable dose was introduced during tolerance induction with interferon gamma (dual SOTI), and its effectiveness was evaluated. The study population comprised 25 IFA patients. Blood samples were taken for analysis, including complete blood count with differential counts of eosinophils, serum total IgE levels, and specific IgE for allergenic foods. Skin prick tests were conducted with the allergens. Oral food challenges were performed to diagnose IFA. Ten patients received dual SOTI, 5 received classic SOTI, 5 received SOTI without interferon gamma (original SOTI), and 5 were not treated (controls). Patients treated with dual SOTI and classic SOTI using interferon gamma became tolerant to the allergenic food. The tolerable dose was introduced successfully in dual SOTI. It was difficult to proceed with the same dosing protocol used for classic SOTI in cases treated with original SOTI. Following dual SOTI, the systemic reaction to oral intake subsided, but the local skin reaction to contact with the allergenic food persisted. Dual SOTI is an improved protocol for SOTI using interferon gamma for IFA.The local skin reaction and systemic reaction to oral intake were dissociated following dual SOTI. In cases of food allergy, tolerance appears to result from desensitization to allergens.

  19. Modulation of interferon-gamma-induced HLA-DR expression on the human keratinocyte cell line SCC-13 by ultraviolet radiation

    International Nuclear Information System (INIS)

    Khan, I.U.; Boehm, K.D.; Elmets, C.A.

    1993-01-01

    Cell surface expression of major histocompatibility determinants on epidermal keratinocytes is a characteristic feature of a number of inflammatory dermatoses and in all likelihood is caused by diffusion of human leukocyte antigen (HLA)-DR-inducing cytokines from cells present in the dermal mononuclear cell infiltrate. Many of these same disorders respond to ultraviolet (UV) radiation phototherapy. Using the human SCC-13 keratinocyte cell line as a model, UV radiation was found to inhibit interferon-gamma-induced HLA-DR expression. Inhibition correlated closely with decreased steady-state levels of HLA-DR mRNA. These findings provide evidence that the therapeutic effect of UV radiation phototherapy may be mediated by its capacity to down-regulate cytokine-induced keratinocyte HLA-DR expression. (Author)

  20. A simple fluorescence based assay for quantification of human immunodeficiency virus particle release

    Directory of Open Access Journals (Sweden)

    Heuser Anke-Mareil

    2010-04-01

    Full Text Available Abstract Background The assembly and release of human immunodeficiency virus (HIV particles from infected cells represent attractive, but not yet exploited targets for antiretroviral therapy. The availability of simple methods to measure the efficiency of these replication steps in tissue culture would facilitate the identification of host factors essential for these processes as well as the screening for lead compounds acting as specific inhibitors of particle formation. We describe here the development of a rapid cell based assay for quantification of human immunodeficiency virus type 1 (HIV-1 particle assembly and/or release. Results Using a fluorescently labelled HIV-derivative, which carries an eYFP domain within the main viral structural protein Gag in the complete viral protein context, the release of virus like particles could be monitored by directly measuring the fluorescence intensity of the tissue culture supernatant. Intracellular Gag was quantitated in parallel by direct fluorescence analysis of cell lysates, allowing us to normalize for Gag expression efficiency. The assay was validated by comparison with p24 capsid ELISA measurements, a standard method for quantifying HIV-1 particles. Optimization of conditions allowed the robust detection of particle amounts corresponding to 50 ng p24/ml in medium by fluorescence spectroscopy. Further adaptation to a multi-well format rendered the assay suitable for medium or high throughput screening of siRNA libraries to identify host cell factors involved in late stages of HIV replication, as well as for random screening approaches to search for potential inhibitors of HIV-1 assembly or release. Conclusions The fast and simple fluorescence based quantification of HIV particle release yielded reproducible results which were comparable to the well established ELISA measurements, while in addition allowing the parallel determination of intracellular Gag expression. The protocols described here

  1. Quality Control Assays for Clinical-Grade Human Mesenchymal Stromal Cells: Methods for ATMP Release.

    Science.gov (United States)

    Radrizzani, Marina; Soncin, Sabrina; Lo Cicero, Viviana; Andriolo, Gabriella; Bolis, Sara; Turchetto, Lucia

    2016-01-01

    Mesenchymal stromal/stem cells (MSC) are promising candidates for the development of cell-based therapies for various diseases and are currently being evaluated in a number of clinical trials (Sharma et al., Transfusion 54:1418-1437, 2014; Ikebe and Suzuki, Biomed Res Int 2014:951512, 2014). MSC for therapeutic applications are classified as advanced therapy medicinal products (ATMP) (Regulation (EC) No 1394/2007 of the European Parliament and of the Council of 13 November 2007 on advanced therapy medicinal products and amending Directive 2001/83/EC and Regulation (EC) No 726/2004) and must be prepared according to good manufacturing practices ( http://ec.europa.eu/health/documents/eudralex/vol-4 ). They may be derived from different starting materials (mainly bone marrow (BM), adipose tissue, or cord blood) and applied as fresh or cryopreserved products, in the autologous as well as an allogeneic context (Sharma et al., Transfusion 54:1418-1437, 2014; Ikebe and Suzuki, Biomed Res Int 2014:951512, 2014; Sensebé and Bourin, Transplantation 87(9 Suppl):S49-S53, 2009). In any case, they require an approved and well-defined panel of assays in order to be released for clinical use.This chapter describes analytical methods implemented and performed in our cell factory as part of the release strategy for an ATMP consisting of frozen autologous BM-derived MSC. Such methods are designed to assess the safety (sterility, endotoxin, and mycoplasma assays) and identity/potency (cell count and viability, immunophenotype and clonogenic assay) of the final product. Some assays are also applied to the biological starting material (sterility) or carried out as in-process controls (sterility, cell count and viability, immunophenotype, clonogenic assay).The validation strategy for each analytical method is described in the accompanying Chapter 20 .

  2. A study of the 51Cr release assay system in micro-method

    International Nuclear Information System (INIS)

    Kiya, Katsuzo; Harada, Kiyoshi; Uozumi, Tohru; Toge, Tetsuya; Hattori, Takao.

    1981-01-01

    Conditions of 51 Cr release assay in microculture were investigated to measure the natural cytotoxic (NC) activity of mouse spleen cells. Malignant glioma (MG) cells of C57BL/6 mouse, induced by 20-methylcholanthrene, were labeled with Na 2 51 CrO 4 . Spleen cells collected from the same mouse strain were suspended in Eagle's MEM. Labeled MG cells and spleen cells were incubated for several hours in a CO 2 incubator. Then the activity of the supernatant was measured by an automatic gamma counter. The optimum conditions of 51 Cr release assay in micro-culture were, (1) number of the target cells: 5 x 10 3 / well (2) FCS concentration: 10% (3) E/T ratio: less than 100 : 1, 50 : 1 was possible (4) incubation time: 15 hours. The number of the target cells at labeling incubation was set to 2 x 10 6 /ml. Though the natural release of 51 Cr was not effected by the viability of the target cells, it was suggested that the NC activity was dependent on the viability of both cells, target and effector cells. (Tsunoda, M.)

  3. Study of the /sup 51/Cr release assay system in micro-method

    Energy Technology Data Exchange (ETDEWEB)

    Kiya, K.; Harada, K.; Uozumi, T. (Hiroshima Univ. (Japan). School of Medicine); Toge, T.; Hattori, T.

    1981-05-01

    Conditions of /sup 51/Cr release assay in microculture were investigated to measure the natural cytotoxic (NC) activity of mouse spleen cells. Malignant glioma (MG) cells of C57BL/6 mouse, induced by 20-methylcholanthrene, were labeled with Na/sub 2//sup 51/CrO/sub 4/. Spleen cells collected from the same mouse strain were suspended in Eagle's MEM. Labeled MG cells and spleen cells were incubated for several hours in a CO/sub 2/ incubator. Then the activity of the supernatant was measured by an automatic gamma counter. The optimum conditions of /sup 51/Cr release assay in micro-culture were, (1) number of the target cells: 5 x 10/sup 3// well (2) FCS concentration: 10% (3) E/T ratio: less than 100 : 1, 50 : 1 was possible (4) incubation time: 15 hours. The number of the target cells at labeling incubation was set to 2 x 10/sup 6//ml. Though the natural release of /sup 51/Cr was not effected by the viability of the target cells, it was suggested that the NC activity was dependent on the viability of both cells, target and effector cells.

  4. Measurement of Nonribosomal Peptide Synthetase Adenylation Domain Activity Using a Continuous Hydroxylamine Release Assay.

    Science.gov (United States)

    Duckworth, Benjamin P; Wilson, Daniel J; Aldrich, Courtney C

    2016-01-01

    Adenylation is a crucial enzymatic process in the biosynthesis of nonribosomal peptide synthetase (NRPS) derived natural products. Adenylation domains are considered the gatekeepers of NRPSs since they select, activate, and load the carboxylic acid substrate onto a downstream peptidyl carrier protein (PCP) domain of the NRPS. We describe a coupled continuous kinetic assay for NRPS adenylation domains that substitutes the PCP domain with hydroxylamine as the acceptor molecule. The pyrophosphate released from the first-half reaction is then measured using a two-enzyme coupling system, which detects conversion of the chromogenic substrate 7-methylthioguanosine (MesG) to 7-methylthioguanine. From profiling substrate specificity of unknown or engineered adenylation domains to studying chemical inhibition of adenylating enzymes, this robust assay will be of widespread utility in the broad field NRPS enzymology.

  5. Calcein AM release-based cytotoxic cell assay for fish leucocytes.

    Science.gov (United States)

    Iwanowicz, Luke R; Densmore, Christine L; Ottinger, Christopher A

    2004-02-01

    A non-specific cytotoxic cell assay for fish is presented that is based on the release of the activated fluorochrome calcein AM from lysed carp epithelioma papulosum cyprini (EPC) cells. To establish the suitability of treating EPC cells with calcein AM the uptake and spontaneous release of the calcein AM by the EPC cells was evaluated. Incubation of 5 microM calcein AM in culture medium with 1x10(5)EPC cells well(-1)for a minimum of 3 h provided sufficient labelling. Spontaneous release of fluorescence from the labelled EPC cells during 10 h of post labelling incubation ranged from 30 to 39% of the total observed fluorescence. Cytotoxic activity of trout leucocytes was evaluated at three leucocyte to target cell ratios (10:1, 2:1 and 1:1) following incubation (4, 6, 8, and 10 h) with calcein AM-labelled EPC cells at 15 degrees C. In some instances, the monoclonal antibody specific for the NCC surface receptor NCCRP-1 (MAb5C.6) was included in the cultures. The activity of NCC cells was significantly inhibited in the presence of 0.25 microg well(-1)of MAb5C.6 relative to no antibody (Prelease assay.

  6. Myxoma virus M-T7, a secreted homolog of the interferon-gamma receptor, is a critical virulence factor for the development of myxomatosis in European rabbits.

    Science.gov (United States)

    Mossman, K; Nation, P; Macen, J; Garbutt, M; Lucas, A; McFadden, G

    1996-01-01

    Myxoma virus is a leporipoxvirus of New World rabbits (Sylvilagus sp.) that induces a rapidly lethal infection known as myxomatosis in the European rabbit (Oryctolagus cuniculus). Like all poxviruses, myxoma virus encodes a plethora of proteins to circumvent or inhibit a variety of host antiviral immune mechanisms. M-T7, the most abundantly secreted protein of myxoma virus-infected cells, was originally identified as an interferon-gamma receptor homolog (Upton, Mossman, and McFadden, Science 258, 1369-1372, 1992). Here, we demonstrate that M-T7 is dispensable for virus replication in cultured cells but is a critical virulence factor for virus pathogenesis in European rabbits. Disruption of both copies of the M-T7 gene in myxoma virus was achieved by the deletion of 372 bp of M-T7 coding sequences, replacement with a selectable marker, p7.5Ecogpt, and selection of a recombinant virus (vMyxlac-T7gpt) resistant to mycophenolic acid. vMyxlac-T7gpt expressed no detectable M-T7 protein and infected cells supernatants were devoid of any detectable interferon-gamma binding activities. Immunohistochemical staining with anti-beta-galactosidase and anti-CD43 antibodies demonstrated that in vMyxlac-T7gpt-infected rabbits the loss of M-T7 not only caused a dramatic reduction in disease symptoms and viral dissemination to secondary sites, but also dramatically influenced host leukocyte behavior. Notably, primary lesions in wild-type virus infections were generally underlayed by large masses of inflammatory cells that did not effectively migrate into the dermal sites of viral replication, whereas in vMyxlac-T7gpt infections this apparent block to leukocyte influx was relieved. A second major phenotypic distinction noted for the M-T7 knockout virus was the extensive activation of lymphocytes in secondary immune organs, particularly the spleen and lymph nodes, by Day 4 of the infection. This is in stark contrast to infection by wild-type myxoma virus, which results in relatively

  7. Automated assay for screening the enzymatic release of reducing sugars from micronized biomass

    Directory of Open Access Journals (Sweden)

    Asther Marcel

    2010-07-01

    Full Text Available Abstract Background To reduce the production cost of bioethanol obtained from fermentation of the sugars provided by degradation of lignocellulosic biomass (i.e., second generation bioethanol, it is necessary to screen for new enzymes endowed with more efficient biomass degrading properties. This demands the set-up of high-throughput screening methods. Several methods have been devised all using microplates in the industrial SBS format. Although this size reduction and standardization has greatly improved the screening process, the published methods comprise one or more manual steps that seriously decrease throughput. Therefore, we worked to devise a screening method devoid of any manual steps. Results We describe a fully automated assay for measuring the amount of reducing sugars released by biomass-degrading enzymes from wheat-straw and spruce. The method comprises two independent and automated steps. The first step is the making of "substrate plates". It consists of filling 96-well microplates with slurry suspensions of micronized substrate which are then stored frozen until use. The second step is an enzymatic activity assay. After thawing, the substrate plates are supplemented by the robot with cell-wall degrading enzymes where necessary, and the whole process from addition of enzymes to quantification of released sugars is autonomously performed by the robot. We describe how critical parameters (amount of substrate, amount of enzyme, incubation duration and temperature were selected to fit with our specific use. The ability of this automated small-scale assay to discriminate among different enzymatic activities was validated using a set of commercial enzymes. Conclusions Using an automatic microplate sealer solved three main problems generally encountered during the set-up of methods for measuring the sugar-releasing activity of plant cell wall-degrading enzymes: throughput, automation, and evaporation losses. In its present set-up, the

  8. Prenatal Dexamethasone and Postnatal High-Fat Diet Decrease Interferon Gamma Production through an Age-Dependent Histone Modification in Male Sprague-Dawley Rats

    Science.gov (United States)

    Yu, Hong-Ren; Tain, You-Lin; Sheen, Jiunn-Ming; Tiao, Mao-Meng; Chen, Chih-Cheng; Kuo, Ho-Chang; Hung, Pi-Lien; Hsieh, Kai-Sheng; Huang, Li-Tung

    2016-01-01

    Overexposure to prenatal glucocorticoid (GC) disturbs hypothalamic-pituitary-adrenocortical axis-associated neuroendocrine metabolism and susceptibility to metabolic syndrome. A high-fat (HF) diet is a major environmental factor that can cause metabolic syndrome. We aimed to investigate whether prenatal GC plus a postnatal HF diet could alter immune programming in rat offspring. Pregnant Sprague-Dawley rats were given intraperitoneal injections of dexamethasone or saline at 14–21 days of gestation. Male offspring were then divided into four groups: vehicle, prenatal dexamethasone exposure, postnatal HF diet (VHF), and prenatal dexamethasone exposure plus a postnatal HF diet (DHF). The rats were sacrificed and adaptive immune function was evaluated. Compared to the vehicle, the DHF group had lower interferon gamma (IFN-γ) production by splenocytes at postnatal day 120. Decreases in H3K9 acetylation and H3K36me3 levels at the IFN-γ promoter correlated with decreased IFN-γ production. The impaired IFN-γ production and aberrant site-specific histone modification at the IFN-γ promoter by prenatal dexamethasone treatment plus a postnatal HF diet resulted in resilience at postnatal day 180. Prenatal dexamethasone and a postnatal HF diet decreased IFN-γ production through a site-specific and an age-dependent histone modification. These findings suggest a mechanism by which prenatal exposure to GC and a postnatal environment exert effects on fetal immunity programming. PMID:27669212

  9. Prenatal Dexamethasone and Postnatal High-Fat Diet Decrease Interferon Gamma Production through an Age-Dependent Histone Modification in Male Sprague-Dawley Rats

    Directory of Open Access Journals (Sweden)

    Hong-Ren Yu

    2016-09-01

    Full Text Available Overexposure to prenatal glucocorticoid (GC disturbs hypothalamic-pituitary-adrenocortical axis-associated neuroendocrine metabolism and susceptibility to metabolic syndrome. A high-fat (HF diet is a major environmental factor that can cause metabolic syndrome. We aimed to investigate whether prenatal GC plus a postnatal HF diet could alter immune programming in rat offspring. Pregnant Sprague-Dawley rats were given intraperitoneal injections of dexamethasone or saline at 14–21 days of gestation. Male offspring were then divided into four groups: vehicle, prenatal dexamethasone exposure, postnatal HF diet (VHF, and prenatal dexamethasone exposure plus a postnatal HF diet (DHF. The rats were sacrificed and adaptive immune function was evaluated. Compared to the vehicle, the DHF group had lower interferon gamma (IFN-γ production by splenocytes at postnatal day 120. Decreases in H3K9 acetylation and H3K36me3 levels at the IFN-γ promoter correlated with decreased IFN-γ production. The impaired IFN-γ production and aberrant site-specific histone modification at the IFN-γ promoter by prenatal dexamethasone treatment plus a postnatal HF diet resulted in resilience at postnatal day 180. Prenatal dexamethasone and a postnatal HF diet decreased IFN-γ production through a site-specific and an age-dependent histone modification. These findings suggest a mechanism by which prenatal exposure to GC and a postnatal environment exert effects on fetal immunity programming.

  10. Continuous in vivo infusion of interferon-gamma (IFN-gamma) enhances engraftment of syngeneic wild-type cells in Fanca-/- and Fancg-/- mice.

    Science.gov (United States)

    Si, Yue; Ciccone, Samantha; Yang, Feng-Chun; Yuan, Jin; Zeng, Daisy; Chen, Shi; van de Vrugt, Henri J; Critser, John; Arwert, Fre; Haneline, Laura S; Clapp, D Wade

    2006-12-15

    Fanconi anemia (FA) is a heterogeneous genetic disorder characterized by bone marrow (BM) failure and cancer susceptibility. Identification of the cDNAs of FA complementation types allows the potential of using gene transfer technology to introduce functional cDNAs as transgenes into autologous stem cells and provide a cure for the BM failure in FA patients. However, strategies to enhance the mobilization, transduction, and engraftment of exogenous stem cells are required to optimize efficacy prior to widespread clinical use. Hypersensitivity of Fancc-/- cells to interferon-gamma (IFN-gamma), a nongenotoxic immune-regulatory cytokine, enhances engraftment of syngeneic wild-type (WT) cells in Fancc-/- mice. However, whether this phenotype is of broad relevance in other FA complementation groups is unresolved. Here we show that primitive and mature myeloid progenitors in Fanca-/- and Fancg-/- mice are hypersensitive to IFN-gamma and that in vivo infusion of IFN-gamma at clinically relevant concentrations was sufficient to allow consistent long-term engraftment of isogenic WT repopulating stem cells. Given that FANCA, FANCC, and FANCG complementation groups account for more than 90% of all FA patients, these data provide evidence that IFN-gamma conditioning may be a useful nongenotoxic strategy for myelopreparation in FA patients.

  11. Continuous in vivo infusion of interferon-gamma (IFN-γ) enhances engraftment of syngeneic wild-type cells in Fanca–/– and Fancg–/– mice

    Science.gov (United States)

    Si, Yue; Ciccone, Samantha; Yang, Feng-Chun; Yuan, Jin; Zeng, Daisy; Chen, Shi; van de Vrugt, Henri J.; Critser, John; Arwert, Fre; Haneline, Laura S.; Clapp, D. Wade

    2006-01-01

    Fanconi anemia (FA) is a heterogeneous genetic disorder characterized by bone marrow (BM) failure and cancer susceptibility. Identification of the cDNAs of FA complementation types allows the potential of using gene transfer technology to introduce functional cDNAs as transgenes into autologous stem cells and provide a cure for the BM failure in FA patients. However, strategies to enhance the mobilization, transduction, and engraftment of exogenous stem cells are required to optimize efficacy prior to widespread clinical use. Hypersensitivity of Fancc–/– cells to interferon-gamma (IFN-γ), a nongenotoxic immune-regulatory cytokine, enhances engraftment of syngeneic wild-type (WT) cells in Fancc–/– mice. However, whether this phenotype is of broad relevance in other FA complementation groups is unresolved. Here we show that primitive and mature myeloid progenitors in Fanca–/– and Fancg–/– mice are hypersensitive to IFN-γ and that in vivo infusion of IFN-γ at clinically relevant concentrations was sufficient to allow consistent long-term engraftment of isogenic WT repopulating stem cells. Given that FANCA, FANCC, and FANCG complementation groups account for more than 90% of all FA patients, these data provide evidence that IFN-γ conditioning may be a useful nongenotoxic strategy for myelopreparation in FA patients. PMID:16946306

  12. Macrophage-Lineage Cells Negatively Regulate the Hematopoietic Stem Cell Pool in Response to Interferon Gamma at Steady State and During Infection.

    Science.gov (United States)

    McCabe, Amanda; Zhang, Yubin; Thai, Vinh; Jones, Maura; Jordan, Michael B; MacNamara, Katherine C

    2015-07-01

    Bone marrow (BM) resident macrophages (Mϕs) regulate hematopoietic stem cell (HSC) mobilization; however, their impact on HSC function has not been investigated. We demonstrate that depletion of BM resident Mϕs increases HSC proliferation as well as the pool of quiescent HSCs. At the same time, during bacterial infection where BM resident Mϕs are selectively increased we observe a decrease in HSC numbers. Moreover, strategies that deplete or reduce Mϕs during infection prevent HSC loss and rescue HSC function. We previously found that the transient loss of HSCs during infection is interferon-gamma (IFNγ)-dependent. We now demonstrate that IFNγ signaling specifically in Mϕs is critical for both the diminished HSC pool and maintenance of BM resident Mϕs during infection. In addition to the IFNγ-dependent loss of BM HSC and progenitor cells (HSPCs) during infection, IFNγ reduced circulating HSPC numbers. Importantly, under infection conditions AMD3100 or G-CSF-induced stem cell mobilization was impaired. Taken together, our data show that IFNγ acts on Mϕs, which are a negative regulator of the HSC pool, to drive the loss in BM and peripheral HSCs during infection. Our findings demonstrate that modulating BM resident Mϕ numbers can impact HSC function in vivo, which may be therapeutically useful for hematologic conditions and refinement of HSC transplantation protocols. © 2015 AlphaMed Press.

  13. Determination of the radionuclide release factor for an evaporator process using nondestructive assay

    International Nuclear Information System (INIS)

    Johnson, R.E.

    1998-01-01

    The 242-A Evaporator is the primary waste evaporator for the Hanford Site radioactive liquid waste stored in underground double-shell tanks. Low pressure evaporation is used to remove water from the waste, thus reducing the amount of tank space required for storage. The process produces a concentrated slurry, a process condensate, and an offgas. The offgas exhausts through two stages of high-efficiency particulate air (HEPA) filters before being discharged to the atmosphere 40 CFR 61 Subpart H requires assessment of the unfiltered exhaust to determine if continuous compliant sampling is required. Because potential (unfiltered) emissions are not measured, methods have been developed to estimate these emissions. One of the methods accepted by the Environmental Protection Agency is the measurement of the accumulation of radionuclides on the HEPA filters. Nondestructive assay (NDA) was selected for determining the accumulation on the HEPA filters. NDA was performed on the HEPA filters before and after a campaign in 1997. NDA results indicate that 2.1 E+4 becquerels of cesium-137 were accumulated on the primary HEPA 1700 filter during the campaign. The feed material processed in the campaign contained a total of 1.4 E+l6 Bq of cesium-137. The release factor for the evaporator process is 1.5 E-12. Based on this release factor, continuous compliant sampling is not required

  14. Antigen-Specific IP-10 Release Is a Sensitive Biomarker of Mycobacterium bovis Infection in Cattle.

    Directory of Open Access Journals (Sweden)

    Sven D C Parsons

    Full Text Available The most widely used ante-mortem diagnostic tests for tuberculosis in cattle are the tuberculin skin test and the interferon-gamma (IFN-γ release assay, both of which measure cell-mediated immune responses to Mycobacterium bovis infection. However, limitations in the performance of these tests results in a failure to identify all infected animals. In attempting to increase the range of diagnostic tests for tuberculosis, measurement of the cytokine IP-10 in antigen-stimulated blood has previously been shown to improve the detection of M. tuberculosis and M. bovis infection, in humans and African buffaloes (Syncerus caffer, respectively. In the present study, 60 cattle were identified by the single intradermal comparative tuberculin test as tuberculosis reactors (n = 24 or non-reactors (n = 36 and the release of IFN-γ and IP-10 in antigen-stimulated whole blood from these animals was measured using bovine specific ELISAs. There was a strong correlation between IP-10 and IFN-γ production in these samples. Moreover, measurement of the differential release of IP-10 in response to stimulation with M. bovis purified protein derivative (PPD and M. avium PPD distinguished between reactor and non-reactor cattle with a sensitivity of 100% (95% CI, 86%-100% and a specificity of 97% (95% CI, 85%-100%. These results suggest that IP-10 might prove valuable as a diagnostic biomarker of M. bovis infection in cattle.

  15. A novel multiparametric flow cytometry-based cytotoxicity assay simultaneously immunophenotypes effector cells: Comparisons to a 4 h 51Cr-release assay

    OpenAIRE

    Kim, GG; Donnenberg, VS; Donnenberg, AD; Gooding, W; Whiteside, TL

    2007-01-01

    Natural killer (NK) cell- or T cell-mediated cytotoxicity traditionally is measured in 4-16h 51Cr-release assays (CRA). A new four-color flow cytometry-based cytotoxicity assay (FCC) was developed to simultaneously measure NK cell cytotoxicity and NK cell phenotype (CD3−CD16+CD56+). Target cells, K562 or Daudi, were labeled with Cell Tracker Orange (CTO) prior to the addition of effector cells. Following co-incubation, 7 amino-actinomycin D (7-AAD) was added to measure death of target cells. ...

  16. Clinical value of whole-blood interferon-gamma assay in patients with suspected pulmonary tuberculosis and AFB smear- and polymerase chain reaction-negative bronchial aspirates.

    Science.gov (United States)

    Lee, Jaehee; Lee, Shin Yup; Yoo, Seung Soo; Cha, Seung Ick; Won, Dong Il; Park, Jae Yong; Lee, Won-Kil; Kim, Chang Ho

    2012-07-01

    Combining a polymerase chain reaction (PCR) test with bronchoscopy is frequently performed to allow a rapid diagnosis of smear-negative pulmonary tuberculosis (PTB). However, limited data are available concerning clinical judgment in patients with suspected PTB and AFB smear- and PCR-negative bronchial aspirates (BA). The present study evaluated the usefulness of whole-blood QuantiFERON-TB Gold In-Tube (QFT) testing in these patients. Of 166 patients with suspected PTB who had undergone bronchoscopy because of smear-negative sputum or inadequate sputum production, 93 (56%) were diagnosed with culture-positive PTB. Seventy-four patients were either AFB smear- or PCR-positive. In the 75 patients whose BA AFB smear and PCR results were both negative, 19 were finally diagnosed with PTB by culture. The QFT test had a negative predictive value of 91% for PTB. The QFT test may be useful for excluding PTB in patients with suspected PTB whose BA AFB smear and PCR results are both negative. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. A novel mechanism of skin tumor promotion involving interferon-gamma (IFNγ)/signal transducer and activator of transcription-1 (Stat1) signaling.

    Science.gov (United States)

    Bozeman, Ronald; Abel, Erika L; Macias, Everardo; Cheng, Tianyi; Beltran, Linda; DiGiovanni, John

    2015-08-01

    The current study was designed to explore the role of signal transducer and activator of transcription 1 (Stat1) during tumor promotion using the mouse skin multistage carcinogenesis model. Topical treatment with both 12-O-tetradecanoylphorbol-13-acetate (TPA) and 3-methyl-1,8-dihydroxy-9-anthrone (chrysarobin or CHRY) led to rapid phosphorylation of Stat1 on both tyrosine (Y701) and serine (S727) residues in epidermis. CHRY treatment also led to upregulation of unphosphorylated Stat1 (uStat1) at later time points. CHRY treatment also led to upregulation of interferon regulatory factor 1 (IRF-1) mRNA and protein, which was dependent on Stat1. Further analyses demonstrated that topical treatment with CHRY but not TPA upregulated interferon-gamma (IFNγ) mRNA in the epidermis and that the induction of both IRF-1 and uStat1 was dependent on IFNγ signaling. Stat1 deficient (Stat1(-/-) ) mice were highly resistant to skin tumor promotion by CHRY. In contrast, the tumor response (in terms of both papillomas and squamous cell carcinomas) was similar in Stat1(-/-) mice and wild-type littermates with TPA as the promoter. Maximal induction of both cyclooxygenase-2 and inducible nitric oxide synthase in epidermis following treatment with CHRY was also dependent on the presence of functional Stat1. These studies define a novel mechanism associated with skin tumor promotion by the anthrone class of tumor promoters involving upregulation of IFNγ signaling in the epidermis and downstream signaling through activated (phosphorylated) Stat1, IRF-1 and uStat1. © 2014 Wiley Periodicals, Inc.

  18. Sphingosine kinase inhibitor suppresses IL-18-induced interferon-gamma production through inhibition of p38 MAPK activation in human NK cells

    International Nuclear Information System (INIS)

    Cheon, Soyoung; Song, Seok Bean; Jung, Minkyung; Park, Yoorim; Bang, Jung-Wook; Kim, Tae Sung; Park, Hyunjeong; Kim, Cherl-hyun; Yang, Yool-hee; Bang, Sa Ik; Cho, Daeho

    2008-01-01

    Natural killer (NK) cells play an important role in the innate immune response. Interleukin-18 (IL-18) is a well-known interferon-gamma (IFN-γ inducing factor, which stimulates immune response in NK and T cells. Sphingosine kinase (SPHK) catalyzes the formation of sphingosine 1-phosphate (S1P), which acts as a second messenger to function as an anti-apoptotic factor and proliferation stimulator of immune cells. In this study, to elucidate whether SPHK is involved in IL-18-induced IFN-γ production, we measured IL-18-induced IFN-γ production after pre-treatment with SPHK inhibitor (SKI) in NK-92MI cells. We found that IL-18-induced IFN-γ expression was blocked by SKI pre-treatment in both mRNA and protein levels. In addition, the increased IFN-γ production by stimulation with IL-18 is mediated through both SPHK and p38 MAPK. To determine the upstream signals of SKI and p38 MAPK in IL-18-induced IFN-γ production, phosphorylation levels of p38 MAPK was measured after SKI pre-treatment. As a result, inhibition of SPHK by SKI blocked phosphorylation of p38 MAPK, showing that SPHK activation by IL-18 is an upstream signal of p38 MAPK activation. Inhibition of SPHK by SKI also inhibited IL-18-induced IFN-γ production in human primary NK cells. In conclusion, SPHK activation is an essential factor for IL-18-induced IFN-γ production via p38 MAPK

  19. ROS mediates interferon gamma induced phosphorylation of Src, through the Raf/ERK pathway, in MCF-7 human breast cancer cell line.

    Science.gov (United States)

    Zibara, Kazem; Zeidan, Asad; Bjeije, Hassan; Kassem, Nouhad; Badran, Bassam; El-Zein, Nabil

    2017-03-01

    Interferon gamma (IFN-ɣ) is a pleiotropic cytokine which plays dual contrasting roles in cancer. Although IFN-ɣ has been clinically used to treat various malignancies, it was recently shown to have protumorigenic activities. Reactive oxygen species (ROS) are overproduced in cancer cells, mainly due to NADPH oxidase activity, which results into several changes in signaling pathways. In this study, we examined IFN-ɣ effect on the phosphorylation levels of key signaling proteins, through ROS production, in the human breast cancer cell line MCF-7. After treatment by IFN-ɣ, results showed a significant increase in the phosphorylation of STAT1, Src, raf, AKT, ERK1/2 and p38 signaling molecules, in a time specific manner. Src and Raf were found to be involved in early stages of IFN-ɣ signaling since their phosphorylation increased very rapidly. Selective inhibition of Src-family kinases resulted in an immediate significant decrease in the phosphorylation status of Raf and ERK1/2, but not p38 and AKT. On the other hand, IFN-ɣ resulted in ROS generation, through H 2 O 2 production, whereas pre-treatment with the ROS inhibitor NAC caused ROS inhibition and a significant decrease in the phosphorylation levels of AKT, ERK1/2, p38 and STAT1. Moreover, pretreatment with a selective NOX1 inhibitor resulted in a significant decrease of AKT phosphorylation. Finally, no direct relationship was found between ROS production and calcium mobilization. In summary, IFN-ɣ signaling in MCF-7 cell line is ROS-dependent and follows the Src/Raf/ERK pathway whereas its signaling through the AKT pathway is highly dependent on NOX1.

  20. Regulation of CD95 expression and CD95-mediated cell death by interferon-gamma in acute lymphoblastic leukemia with chromosomal translocation t(4;11).

    Science.gov (United States)

    Dörrie, J; Schuh, W; Keil, A; Bongards, E; Greil, J; Fey, G H; Zunino, S J

    1999-10-01

    The regulatory effects of IFNgamma on CD95 expression and CD95-mediated cell death were investigated in three high-risk pro-B acute lymphoblastic leukemia (ALL) lines that carry the chromosomal translocation t(4;11)(q21;q23). These leukemias are characteristically refractory to conventional chemotherapeutic treatments operating through the induction of apoptosis. However, the mechanisms leading to increased cell survival and resistance to cell death in these leukemias are largely unknown. Interferon-gamma (IFNgamma), a potent inhibitor of hematopoiesis, acts in part by upregulating CD95 and sensitizing cells to CD95-induced apoptosis. The t(4;11) lines SEM, RS4;11, and MV4;11 expressed low levels of CD95, but were completely resistant to CD95-mediated death. Addition of IFNgamma markedly upregulated CD95 expression in SEM (8-9-fold), RS4;11 (2-3-fold), and MV4;11 (2-3-fold) lines. However, after treatment with IFNgamma, only an 11% increase in sensitivity to CD95-mediated cell death was observed in SEM cells, whereas RS4;11 and MV4;11 cells remained resistant. Cycloheximide, but not actinomycin D or brefeldin A, increased CD95-specific cell death only in IFNgamma-treated RS4;11 cells by approximately 12%. Abundant levels of Bcl-2 and Bcl-XL, known to inhibit CD95-signaling in some cells, were present suggesting a possible role for both molecules in the resistance to CD95-mediated cell death. Resistance of the leukemic blasts to CD95-mediated cell death and the failure of IFNgamma to substantially sensitize the CD95-signaling pathway may contribute to the highly malignant phenotype of pro-B ALL with translocation t(4;11).

  1. Proinflammatory cytokines tumor necrosis factor-alpha and interferon-gamma alter tight junction structure and function in the rat parotid gland Par-C10 cell line.

    Science.gov (United States)

    Baker, Olga J; Camden, Jean M; Redman, Robert S; Jones, Jonathan E; Seye, Cheikh I; Erb, Laurie; Weisman, Gary A

    2008-11-01

    Sjögren's syndrome (SS) is an autoimmune disorder characterized by inflammation and dysfunction of salivary glands, resulting in impaired secretory function. The production of the proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) is elevated in exocrine glands of patients with SS, although little is known about the effects of these cytokines on salivary epithelial cell functions necessary for saliva secretion, including tight junction (TJ) integrity and the establishment of transepithelial ion gradients. The present study demonstrates that chronic exposure of polarized rat parotid gland (Par-C10) epithelial cell monolayers to TNF-alpha and IFN-gamma decreases transepithelial resistance (TER) and anion secretion, as measured by changes in short-circuit current (I(sc)) induced by carbachol, a muscarinic cholinergic receptor agonist, or UTP, a P2Y(2) nucleotide receptor agonist. In contrast, TNF-alpha and IFN-gamma had no effect on agonist-induced increases in the intracellular calcium concentration [Ca(2+)](i) in Par-C10 cells. Furthermore, treatment of Par-C10 cell monolayers with TNF-alpha and IFN-gamma increased paracellular permeability to normally impermeant proteins, altered cell and TJ morphology, and downregulated the expression of the TJ protein, claudin-1, but not other TJ proteins expressed in Par-C10 cells. The decreases in TER, agonist-induced transepithelial anion secretion, and claudin-1 expression caused by TNF-alpha, but not IFN-gamma, were reversible by incubation of Par-C10 cell monolayers with cytokine-free medium for 24 h, indicating that IFN-gamma causes irreversible inhibition of cellular activities associated with fluid secretion in salivary glands. Our results suggest that cytokine production is an important contributor to secretory dysfunction in SS by disrupting TJ integrity of salivary epithelium.

  2. Studies of methotrexate-induced limb dysplasias utilizing a 51chromium release assay

    International Nuclear Information System (INIS)

    Brewton, R.G.; MacCabe, J.A.

    1990-01-01

    The folate antagonist methotrexate (MTX), widely used in chemotherapy, is a well-documented teratogen. However, the mechanism by which it exerts its effects is still unclear. Specifically, we have examined the cytotoxicity of MTX in vivo and in vitro and have looked at the relationship between cytotoxicity and teratogenesis. The chick embryo was utilized to examine the effects of the drug administered to carefully staged embryos. Embryos were exposed at stages 18-22 and examined on day 11 of incubation. Wings were malformed in a stage-dependent manner while legs were affected similarly at each stage used. A modification of the 51chromium-release assay was used to test the toxicity of MTX to limb cells in vitro. None of the tissues tested showed measurable toxicity in vitro even though the drug kills cells in vivo, thereby suggesting that MTX may be metabolized differently in vitro. Malformations induced by MTX do not seem to be due to changes in the amount of cell death taking place in the limb but may be caused by a transient inhibition of cell division

  3. Oleylphosphocholine (OlPC) arrests Cryptosporidium parvum growth in vitro and prevents lethal infection in interferon gamma receptor knock-out mice.

    Science.gov (United States)

    Sonzogni-Desautels, Karine; Renteria, Axel E; Camargo, Fabio V; Di Lenardo, Thomas Z; Mikhail, Alexandre; Arrowood, Michael J; Fortin, Anny; Ndao, Momar

    2015-01-01

    Cryptosporidium parvum is a species of protozoa that causes cryptosporidiosis, an intestinal disease affecting many mammals including humans. Typically, in healthy individuals, cryptosporidiosis is a self-limiting disease. However, C. parvum can cause a severe and persistent infection that can be life-threatening for immunocompromised individuals, such as AIDS patients. As there are no available treatments for these patients that can cure the disease, there is an urgent need to identify treatment options. We tested the anti-parasitic activity of the alkylphosphocholine oleylphosphocholine (OlPC), an analog of miltefosine, against C. parvum in in vitro and in vivo studies. In vitro experiments using C. parvum infected human ileocecal adenocarcinoma cells (HCT-8 cells) showed that OlPC has an EC50 of 18.84 nM. Moreover, no cell toxicity has been seen at concentrations ≤50 μM. C57BL/6 interferon gamma receptor knock-out mice, were infected by gavage with 4000 C. parvum oocysts on Day 0. Oral treatments, with OlPC, miltefosine, paromomycin or PBS, began on Day 3 post-infection for 10 days. Treatment with OlPC, at 40 mg/kg/day resulted in 100% survival, complete clearance of parasite in stools and a 99.9% parasite burden reduction in the intestines at Day 30. Doses of 30 and 20 mg/kg/day also demonstrated an increased survival rate and a dose-dependent parasite burden reduction. Mice treated with 10 mg/kg/day of miltefosine resulted in 50% survival at Day 30. In contrast, control mice, treated with PBS or 100 mg/kg/day of paromomycin, died or had to be euthanized between Days 6 and 13 due to severe illness. Results of parasite burden were obtained by qPCR and cross-validated by both flow cytometry of stool oocysts and histological sections of the ileum. Together, our results strongly support that OlPC represents a potential candidate for the treatment of C. parvum infections in immunocompromised patients.

  4. Oleylphosphocholine (OlPC arrests Cryptosporidium parvum growth in vitro and prevents lethal infection in interferon gamma receptor knock-out mice

    Directory of Open Access Journals (Sweden)

    Karine eSonzogni-Desautels

    2015-09-01

    Full Text Available Cryptosporidium parvum is a species of protozoa that causes cryptosporidiosis, an intestinal disease affecting many mammals including humans. Typically, in healthy individuals, cryptosporidiosis is a self-limiting disease. However, C. parvum can cause a severe and persistent infection that can be life-threatening for immunocompromised individuals, such as AIDS patients. As there are no available treatments for these patients that can cure the disease, there is an urgent need to identify treatment options. We tested the anti-parasitic activity of the alkylphosphocholine oleylphosphocholine (OlPC, an analog of miltefosine, against C. parvum in in vitro and in vivo studies. In vitro experiments using C. parvum infected human ileocecal adenocarcinoma cells (HCT-8 cells showed that OlPC has an EC50 of 18.84 nM. Moreover, no cell toxicity has been seen at concentrations ≤50 µM. C57BL/6 interferon gamma receptor knock-out mice, were infected by gavage with 4000 C. parvum oocysts on Day 0. Oral treatments, with OlPC, miltefosine, paromomycin or PBS, began on Day 3 post-infection for 10 days. Treatment with OlPC, at 40 mg/kg/day resulted in 100% survival, complete clearance of parasite in stools and a 99.9% parasite burden reduction in the intestines at Day 30. Doses of 30 mg/kg/day and 20 mg/kg/day also demonstrated an increased survival rate and a dose-dependent parasite burden reduction. Mice treated with 10 mg/kg/day of miltefosine resulted in 50% survival at Day 30. In contrast, control mice, treated with PBS or 100 mg/kg/day of paromomycin, died or had to be euthanized between Days 6 and 13 due to severe illness. Results of parasite burden were obtained by qPCR and cross-validated by both flow cytometry of stool oocysts and histological sections of the ileum. Together, our results strongly support that OlPC represents a potential candidate for the treatment of C. parvum infections in immunocompromised patients.

  5. A novel multiparametric flow cytometry-based cytotoxicity assay simultaneously immunophenotypes effector cells: comparisons to a 4 h 51Cr-release assay.

    Science.gov (United States)

    Kim, G G; Donnenberg, V S; Donnenberg, A D; Gooding, W; Whiteside, T L

    2007-08-31

    Natural killer (NK) cell-or T cell-mediated cytotoxicity traditionally is measured in 4-16 h (51)Cr-release assays (CRA). A new four-color flow cytometry-based cytotoxicity assay (FCC) was developed to simultaneously measure NK cell cytotoxicity and NK cell phenotype (CD3(-)CD16(+)CD56(+)). Target cells, K562 or Daudi, were labeled with Cell Tracker Orange (CTO) prior to the addition of effector cells. Following co-incubation, 7 amino-actinomycin D (7-AAD) was added to measure death of target cells. The phenotype of effectors, viability of targets, the formation of tumor-effector cell conjugates and absolute numbers of all cells were measured based on light scatter (FSC/SSC), double discrimination of the fluorescence peak integral and height, and fluorescence intensity. Kinetic studies (0.5 and 1 to 4 h) at different effector to target (E:T) cell ratios (50, 25, 12, and 6) confirmed that the 3 h incubation was optimal. The FCC assay is more sensitive than the CRA, has a coefficient of variation (CV) 8-13% and reliably measures NK cell-or lymphokine-activated killer (LAK) cell-mediated killing of target cells in normal controls and subjects with cancer. The FCC assay can be used to study a range of phenotypic attributes, in addition to lytic activity of various subsets of effector cells, without radioactive tracers and thus, it is relatively inexpensive. The FCC assay has a potential for providing information about molecular interactions underlying target cell lysis and thus becoming a major tool for studies of disease pathogenesis as well as development of novel immune therapies.

  6. Martian Superoxide and Peroxide O2 Release (OR) Assay: A New Technology for Terrestrial and Planetary Applications

    Science.gov (United States)

    Georgiou, Christos D.; Zisimopoulos, Dimitrios; Panagiotidis, Konstantinos; Grintzalis, Kontantinos; Papapostolou, Ioannis; Quinn, Richard C.; McKay, Christopher P.; Sun, Henry J.

    2015-01-01

    This study presents an assay for the detection and quantification of soil metal superoxides and peroxides in regolith and soil. The O2 release (OR) assay is based on the enzymatic conversion of the hydrolysis products of metal oxides to O2, and their quantification by an O2 electrode based on the stoichiometry of the involved reactions: The intermediate product O2 from the hydrolysis of metal superoxides is converted by cytochrome c to O2, and also by superoxide dismutase (SOD) to 1/2 mol O2 and 1/2 mol H2O2, which is then converted by catalase (CAT) to 1/2 mol O2. The product H2O2 from the hydrolysis of metal peroxides and hydroperoxides is converted to 1/2 mol O2 by CAT. The assay-method was validated in a sealed sample chamber using a liquid-phase Clark-type O2 electrode with known concentrations of O2 and H2O2, and with commercial metal superoxide and peroxide mixed with Mars analogue Mojave and Atacama Desert soils. Carbonates and perchlorates, both present on Mars, do not interfere with the assay. The assay lower limit of detection, using luminescence quenching/optical sensing O2-electrodes, is 1 nmol O2 cm(exp. -3) or better. The activity of the assay enzymes SOD and cytochrome c was unaffected up to 6 Gy exposure by gamma-radiation, while CAT retained 100% and 40% of its activity at 3 and 6 Gy, respectively, demonstrating the suitability of these enzymes for planetary missions, e.g., in Mars or Europa.

  7. Cell-mediated cytotoxicity for melanome tumor cells: detection by a (3H)proline release assay

    International Nuclear Information System (INIS)

    Saal, J.G.; Rieber, E.P.; Riethmueller, G.

    1976-01-01

    An in vitro lymphocyte-mediated cytotoxicity assay using [ 3 H]proline-labelled target cells is described. The assay, modified from an original procedure of Bean et al., assesses the release of [ 3 H]proline by filtering the total culture fluid containing both trypsinised tumor cells and effector cells. Filtration is performed with a semiautomatic harvesting device using low suction pressure and large-diameter glass filters. Pretreatment of filters with whole serum diminishes adsorption of cell-free radioactive material considerably and thus increases the sensitivity of the assay. Nearly 100% of the radioactivity could be recovered with this harvesting device. The technique allowed the detection of cytolytic activities of lymphocytes after 6 h of incubation. Lymphocytes from patients with primary malignant melanoma showed a significantly higher cytolytic reactivity (p > 0.001) than normal donors' lymphocytes against three different melanoma cell lines. In a series of parallel experiments on 36 patients and 18 normal donors, this modification of the [ 3 ]proline test was compared with three different assays: the conventional microcytotoxicity test of Takasugi and Klein, the original [ 3 H]proline microcytotoxicity test of Bean et al., and the viability count of tumor cells. (Auth.)

  8. Biosensor cell assay for measuring real-time aldosterone-induced release of histamine from mesenteric arteries

    DEFF Research Database (Denmark)

    Dalgaard, Emil G; Andersen, Kenneth; Svenningsen, Per

    2017-01-01

    as a sensitive biosensor assay for histamine release from isolated mouse mesenteric arteries. Activation of the H1 receptor by histamine was measured as an increased number of intracellular Ca(2+) transient peaks using fluorescence imaging RESULTS: The developed biosensor was sensitive to histamine...... in physiological relevant concentrations and responded to substances released by the artery preparation. Aldosterone treatment of mesenteric arteries from wild type mice for 50 minutes resulted in an increased number of intracellular Ca(2+) transient peaks in the biosensor cells, which was significantly inhibited...... by the histamine H1 blocker pyrilamine. Mesenteric arteries from mast cell deficient SASH mice induced similar pyrilamine-sensitive Ca(2+) transient response in the biosensor cells. Mesenteric arteries from wild type and SASH mice expressed histamine decarboxylase mRNA, indicating that mast cells are not the only...

  9. The Poly-γ-D-Glutamic Acid Capsule of Bacillus licheniformis, a Surrogate of Bacillus anthracis Capsule Induces Interferon-Gamma Production in NK Cells through Interactions with Macrophages.

    Science.gov (United States)

    Lee, Hae-Ri; Jeon, Jun Ho; Rhie, Gi-Eun

    2017-05-28

    The poly-γ- D -glutamic acid (PGA) capsule, a major virulence factor of Bacillus anthracis , provides protection of the bacterium from phagocytosis and allows its unimpeded growth in the host. We investigated crosstalk between murine natural killer (NK) cells and macrophages stimulated with the PGA capsule of Bacillus licheniformis , a surrogate of the B. anthracis capsule. PGA induced interferon-gamma production from NK cells cultured with macrophages. This effect was dependent on macrophage-derived IL-12 and cell-cell contact interaction with macrophages through NK cell receptor NKG2D and its ligand RAE-1. The results showed that PGA could enhance NK cell activation by inducing IL-12 production in macrophages and a contact-dependent crosstalk with macrophages.

  10. A high-performance liquid chromatography method for the serotonin release assay is equivalent to the radioactive method.

    Science.gov (United States)

    Sono-Koree, N K; Crist, R A; Frank, E L; Rodgers, G M; Smock, K J

    2016-02-01

    The serotonin release assay (SRA) is considered the gold standard laboratory test for heparin-induced thrombocytopenia (HIT). The historic SRA method uses platelets loaded with radiolabeled serotonin to evaluate platelet activation by HIT immune complexes. However, a nonradioactive method is desirable. We report the performance characteristics of a high-performance liquid chromatography (HPLC) SRA method. We validated the performance characteristics of an HPLC-SRA method, including correlation with a reference laboratory using the radioactive method. Serotonin released from reagent platelets was quantified by HPLC using fluorescent detection. Results were expressed as % release and classified as positive, negative, or indeterminate based on previously published cutoffs. Serum samples from 250 subjects with suspected HIT were tested in the HPLC-SRA and with the radioactive method. Concordant classifications were observed in 230 samples (92%). Sera from 41 healthy individuals tested negative. Between-run imprecision studies showed standard deviation of performance characteristics, equivalent to the historic radioactive method, but avoids the complexities of working with radioactivity. © 2015 John Wiley & Sons Ltd.

  11. Identifying Predictors of Interferon-γ Release Assay Results in Pediatric Latent Tuberculosis: A Protective Role of Bacillus Calmette-Guérin?

    Science.gov (United States)

    Sotgiu, Giovanni; Altet-Gómez, Neus; Tsolia, Maria; Ruga, Ezia; Velizarova, Svetlana; Kampmann, Beate

    2012-01-01

    Rationale: Interferon-γ (IFN-γ) release assays are widely used to diagnose latent infection with Mycobacterium tuberculosis in adults, but their performance in children remains incompletely evaluated to date. Objectives: To investigate factors influencing results of IFN-γ release assays in children using a large European data set. Methods: The Pediatric Tuberculosis Network European Trials group pooled and analyzed data from five sites across Europe comprising 1,128 children who were all investigated for latent tuberculosis infection by tuberculin skin test and at least one IFN-γ release assay. Multivariate analyses examined age, bacillus Calmette-Guérin (BCG) vaccination status, and sex as predictor variables of results. Subgroup analyses included children who were household contacts. Measurements and Main Results: A total of 1,093 children had a QuantiFERON-TB Gold In-Tube assay and 382 had a T-SPOT.TB IFN-γ release assay. Age was positively correlated with a positive blood result (QuantiFERON-TB Gold In-Tube: odds ratio [OR], 1.08 per year increasing age [P 5 yr). Conclusions: Our data show that BCG vaccination may be effective in protecting children against Mycobacterium tuberculosis infection. To restrict use of IFN-γ release assays to children with positive skin tests risks underestimating latent infection. PMID:22700862

  12. Differences in immunoglobulin subclasses between dye exclusion and 51Cr release complement-dependent lymphocytotoxicity assays

    International Nuclear Information System (INIS)

    McConnachie, P.R.; Denegri, J.F.; Lower, F.E.; Torry, D.S.; McIntyre, J.A.

    1986-01-01

    Paradoxical differences previously noted between lymphocytotoxicity detected by dye exclusion at room temperature (CDCE) or by 51 Cr release (CDC 51 Cr) at 37 0 C in maternal antipaternal complement-dependent lymphocytotoxicity have suggested that CDCE and CDC 51 Cr at 37 0 C, but not at 20 degrees C, may detect different immunological antibody-antigen interactions. Reactions in the two test systems against the same target cells were compared in sera from known immune dialysis patients, secondary aborting women, and refractory platelet recipients before and after heat treatment of sera, absorption with solid-phase heparin, anti-light-chain augmentation, and the addition of murine monoclonal anti-IgG subclass antibodies. The results demonstrate significant differences between the two tests using the same target and sera. Further, the results imply the presence of an inhibitor and an inhibitor of inhibitor in sera. The involvement of different immunoglobulin subclasses was shown in the two tests. These data demonstrate the necessity for further study of the nature of the differences in the mechanisms of these clinically important antibody-detecting systems

  13. Direct measurement of human plasma corticotropin-releasing hormone by two-site immunoradiometric assay

    International Nuclear Information System (INIS)

    Linton, E.A.; McLean, C.; Nieuwenhuyzen Kruseman, A.C.; Tilders, F.J.; Van der Veen, E.A.; Lowry, P.J.

    1987-01-01

    A ''two-site'' immunoradiometric assay (IRMA) which allows the direct estimation of human CRH (hCRH) in plasma is described. Using this IRMA, basal levels of CRH in normal subjects ranged from 2-28 pg/mL [mean, 15 +/- 7 (+/- SD) pg/mL; n = 58]. Values in men and women were similar. Plasma CRH values within this range were also found in patients with Cushing's syndrome, Addison's disease, and Nelson's syndrome, with no correlation between plasma CRH and ACTH levels in these patients. Elevated plasma CRH levels were found in pregnant women near term [1462 +/- 752 (+/- SD) pg/mL; n = 55], and the dilution curve of this CRH-like immunoreactivity paralleled the IRMA standard curve. After its immunoadsorption from maternal plasma, this CRH-like material eluted on reverse phase high performance liquid chromatography with a retention time identical to that of synthetic CRH and had equipotent bioactivity with the synthetic peptide in the perfused anterior pituitary cell bioassay. Circulating CRH was not detected in Wistar rats, even after adrenalectomy and subsequent ether stress. Synthetic hCRH was degraded by fresh human plasma relatively slowly; 65% of added CRH remained after 1 h of incubation at 37 C. Degradation was inhibited by heat treatment (54 C; 1 h), cold treatment (4 C; 4 h), or freezing and thawing. Loss of synthetic rat CRH occurred more rapidly when fresh rat plasma was used; only 20% of added CRH remained under the same conditions. The inability to measure CRH in peripheral rat plasma may be due to the presence of active CRH-degrading enzymes which fragment the CRH molecule into forms not recognized by the CRH IRMA

  14. Comparison of two insulin assays for first-phase insulin release in type 1 diabetes prediction and prevention studies.

    Science.gov (United States)

    Mahon, Jeffrey L; Beam, Craig A; Marcovina, Santica M; Boulware, David C; Palmer, Jerry P; Winter, William E; Skyler, Jay S; Krischer, Jeffrey P

    2011-11-20

    Detection of below-threshold first-phase insulin release or FPIR (1+3 minute insulin concentrations during an intravenous glucose tolerance test [IVGTT]) is important in type 1 diabetes prediction and prevention studies including the TrialNet Oral Insulin Prevention Trial. We assessed whether an insulin immunoenzymometric assay (IEMA) could replace the less practical but current standard of a radioimmunoassay (RIA) for FPIR. One hundred thirty-three islet autoantibody positive relatives of persons with type 1 diabetes underwent 161 IVGTTs. Insulin concentrations were measured by both assays in 1056 paired samples. A rule classifying FPIR (below-threshold, above-threshold, uncertain) by the IEMA was derived and validated against FPIR by the RIA. The insulin IEMA-based rule accurately classified below- and above-threshold FPIRs by the RIA in 110/161 (68%) IVGTTs, but was uncertain in 51/161 (32%) tests for which FPIR by RIA is needed. An uncertain FPIR by the IEMA was more likely among below-threshold vs above-threshold FPIRs by the RIA (64% [30/47] vs. 18% [21/114], respectively; pTrialNet is limiting the insulin RIA for FPIR to the latter given the practical advantages of the more specific IEMA. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. Spironolactone inhibits production of proinflammatory cytokines, including tumour necrosis factor-alpha and interferon-gamma, and has potential in the treatment of arthritis

    DEFF Research Database (Denmark)

    Bendtzen, K; Hansen, P R; Rieneck, K

    2003-01-01

    and interleukin 6 (70-90% inhibition). Release of these cytokines was also suppressed when testing whole blood from RA patients receiving 50 mg spironolactone twice daily, indicating that pharmaceutical use of the drug may suppress the release of inflammatory cytokines. Spironolactone therapy was generally well...

  16. C-Reactive Protein (CRP), Interferon Gamma-Inducible Protein 10 (IP-10), and Lipopolysaccharide (LPS) Are Associated with Risk of Tuberculosis after Initiation of Antiretroviral Therapy in Resource-Limited Settings

    Science.gov (United States)

    Tenforde, Mark W.; Gupte, Nikhil; Dowdy, David W.; Asmuth, David M.; Balagopal, Ashwin; Pollard, Richard B.; Sugandhavesa, Patcharaphan; Lama, Javier R.; Pillay, Sandy; Cardoso, Sandra W.; Pawar, Jyoti; Santos, Breno; Riviere, Cynthia; Mwelase, Noluthando; Kanyama, Cecilia; Kumwenda, Johnstone; Hakim, James G.; Kumarasamy, Nagalingeswaran; Bollinger, Robert; Semba, Richard D.; Campbell, Thomas B.; Gupta, Amita

    2015-01-01

    Objective The association between pre-antiretroviral (ART) inflammation and immune activation and risk for incident tuberculosis (TB) after ART initiation among adults is uncertain. Design Nested case-control study (n = 332) within ACTG PEARLS trial of three ART regimens among 1571 HIV-infected, treatment-naïve adults in 9 countries. We compared cases (participants with incident TB diagnosed by 96 weeks) to a random sample of controls (participants who did not develop TB, stratified by country and treatment arm). Methods We measured pre-ART C-reactive protein (CRP), EndoCab IgM, ferritin, interferon gamma (IFN-γ), interleukin 6 (IL-6), interferon gamma-inducible protein 10 (IP-10), lipopolysaccharide (LPS), soluble CD14 (sCD14), tumor necrosis factor alpha (TNF-α), and CD4/DR+/38+ and CD8/DR+/38+ T cells. Markers were defined according to established cutoff definitions when available, 75th percentile of measured values when not, and detectable versus undetectable for LPS. Using logistic regression, we measured associations between biomarkers and incident TB, adjusting for age, sex, study site, treatment arm, baseline CD4 and log10 viral load. We assessed the discriminatory value of biomarkers using receiver operating characteristic (ROC) analysis. Results Seventy-seven persons (4.9%) developed incident TB during follow-up. Elevated baseline CRP (aOR 3.25, 95% CI: 1.55–6.81) and IP-10 (aOR 1.89, 95% CI: 1.05–3.39), detectable plasma LPS (aOR 2.39, 95% CI: 1.13–5.06), and the established TB risk factors anemia and hypoalbuminemia were independently associated with incident TB. In ROC analysis, CRP, albumin, and LPS improved discrimination only modestly for TB risk when added to baseline routine patient characteristics including CD4 count, body mass index, and prior TB. Conclusion Incident TB occurs commonly after ART initiation. Although associated with higher post-ART TB risk, baseline CRP, IP-10, and LPS add limited value to routine patient characteristics

  17. IFN-γ release assay conversions and reversions. Challenges with serial testing in U.S. health care workers.

    Science.gov (United States)

    Joshi, Manish; Monson, Thomas P; Joshi, Anita; Woods, Gail L

    2014-03-01

    IFN-γ release assays (IGRAs) including the QuantiFERON-TB gold in-tube test (QFT-GIT) are increasingly used in place of the tuberculin skin test (TST) in surveillance programs for Mycobacterium tuberculosis infection in the United States. However, data on conversions, reversions, and predictive value of QFT in such programs for health care workers (HCWs) are limited. The purpose of this study is to assess long-term reproducibility and conversion and reversion rates of QFT-GIT among HCWs who underwent serial testing at a tertiary care center in the United States. Retrospective chart review of HCWs at the Central Arkansas Veterans Healthcare System (CAVHS) who underwent serial testing with QFT-GIT as a part of their employee screening between November 1, 2008 and January 31, 2011. A total of 2,303 HCWs had at least 2 QFT-GITs 1 year apart. The initial QFT-GIT was positive for 69 and 2 were indeterminate. Of these 69 HCWs, 31 (45%) reverted on repeat testing, and 25 of 31 (80.6%) HCWs who reverted had a negative look-back TST. Of the 2,232 HCWs with an initial negative QFT-GIT, 71 (3.2%) converted on repeat testing. A third QFT-GIT assay was performed in 41 of the 71 converters and 90% (37 of 41) reverted back to negative. Only two HCWs had TST and QFT-GIT conversion. Poor IGRA reproducibility and a low predictive value of QFT-GIT conversions indicate that QFT-GIT with current interpretation criteria should not be used for serial screening of U.S. HCWs. Negative TSTs have higher reproducibility than QFT-GIT for serial testing of HCWs in low tuberculosis incidence settings.

  18. A randomized, double-blind, phase I/II trial of tumor necrosis factor and interferon-gamma for treatment of AIDS-related complex (Protocol 025 from the AIDS Clinical Trials Group).

    Science.gov (United States)

    Agosti, J M; Coombs, R W; Collier, A C; Paradise, M A; Benedetti, J K; Jaffe, H S; Corey, L

    1992-05-01

    To determine safety and efficacy of tumor necrosis factor (TNF) and interferon-gamma (IFN gamma) in the treatment of patients with acquired immunodeficiency syndrome (AIDS)-related complex, a randomized, double-blind study was conducted. Twenty-five patients with AIDS-related complex and CD4 lymphocytes less than or equal to 500 x 10(6)/L attended an AIDS Clinical Trials Unit of a tertiary referral center. Patients were administered tumor necrosis factor (TNF) (10 micrograms/m2) or IFN gamma (10 micrograms/m2), or both intramuscularly three times weekly for 16 weeks. Side effects from all three preparations included fever, constitutional symptoms, and local reactions. No significant hematologic, hepatic, renal, or coagulation abnormalities were observed. CD4 lymphocyte counts, beta 2-microglobulin, p24 antigen levels, and anti-p24 antibody did not change significantly during therapy. Similarly, no significant change was noted in rates of HIV isolation from peripheral blood mononuclear cells or plasma. TNF and IFN gamma were tolerable after premedication with acetaminophen; however, no significant change in markers of human immunodeficiency virus infection was demonstrated. These cytokines alone do not appear to be of benefit, nor do they appear to hasten the progression of HIV infection.

  19. Cytokine Release Assays as Tests for Exposure to Leishmania, and for Confirming Cure from Leishmaniasis, in Solid Organ Transplant Recipients.

    Directory of Open Access Journals (Sweden)

    Eugenia Carrillo

    Full Text Available Spain has one of the world's largest pools of organ donors and is a global leader in terms of the number of transplants it performs. The current outbreak of leishmaniasis in Fuenlabrada (in the southwest of the region of Madrid, Spain has involved 600 clinical cases since late 2009 (prevalence 0.2%. It may therefore be wise to monitor the town's transplanted population for Leishmania infantum; its members are immunosuppressed and at greater risk of infection and relapse following treatment. The present work examines the use of cytokine release assays to determine the prevalence of Leishmania infection in this population, and to confirm recovery following treatment for visceral leishmaniasis (VL. The humoral and cellular immune responses to L. infantum were characterized in 63 solid organ transplant (SOT recipients from Fuenlabrada, 57 of whom reported no previous episode of VL (NVL subjects, and six of whom had been cured of VL (CVL subjects. Seventeen subjects (12 NVL and 5 CVL showed a patent lymphoproliferative response to soluble Leishmania antigen (SLA. Stimulation of peripheral blood mononuclear cell cultures and of whole blood with SLA led to the production of different combinations of cytokines that might serve to confirm Leishmania infection or recovery from VL and help prevent cured patients from relapsing into this serious condition.

  20. Clinical evaluation of thyrotropin-releasing hormone (TRH) test with a sensitive immunoradiometric thyrotropin (TSH) assay kit

    International Nuclear Information System (INIS)

    Nakamura, Saeko; Demura, Reiko; Yamanaka, Yukako; Ishiwatari, Naoko; Jibiki, Kazuko; Odagiri, Emi; Demura, Hiroshi

    1987-01-01

    Thyrotropin-releasing hormone (TRH) test was performed using a commercially available immunoradiometric thyrotropin (TSH) assay kit (RIA-gnost hTSH) in patients with endocrine diseases. The basal serum concentration of TSH ranged from 0.2 to 2.9 μU/ml in healthy subjects. The values for endocrine diseases, except for Graves' disease, were almost within the normal range. A significant increase in TSH values caused by TRH test was observed in females compared with males (4.4 - 24.7 μU/ml vs 4.1 - 12.3 μU/ml). In cases of Graves' disease, there was a good correlation between the basal TSH value and the response of TSH to TRH. However, in the other endocrine diseases, including acromegaly, prolactinoma, anorexia nervosa, Cushing syndrome, and hypopituitarism, the response of TSH to TRH did not necessarily correlated with the basal TSH value. TRH test would be of value in elucidating pathophysiologic features, as well as in accurately diagnosing secretion reserve of TSH. (Namekawa, K.)

  1. Clinical evaluation of thyrotropin-releasing hormone (TRH) test with a sensitive immunoradiometric thyrotropin (TSH) assay kit

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, Saeko; Demura, Reiko; Yamanaka, Yukako; Ishiwatari, Naoko; Jibiki, Kazuko; Odagiri, Emi; Demura, Hiroshi

    1987-10-01

    Thyrotropin-releasing hormone (TRH) test was performed using a commercially available immunoradiometric thyrotropin (TSH) assay kit (RIA-gnost hTSH) in patients with endocrine diseases. The basal serum concentration of TSH ranged from 0.2 to 2.9 ..mu..U/ml in healthy subjects. The values for endocrine diseases, except for Graves' disease, were almost within the normal range. A significant increase in TSH values caused by TRH test was observed in females compared with males (4.4 - 24.7 ..mu..U/ml vs 4.1 - 12.3 ..mu..U/ml). In cases of Graves' disease, there was a good correlation between the basal TSH value and the response of TSH to TRH. However, in the other endocrine diseases, including acromegaly, prolactinoma, anorexia nervosa, Cushing syndrome, and hypopituitarism, the response of TSH to TRH did not necessarily correlated with the basal TSH value. TRH test would be of value in elucidating pathophysiologic features, as well as in accurately diagnosing secretion reserve of TSH. (Namekawa, K.).

  2. Cost-effectiveness of interferon-γ release assay versus chest X-ray for tuberculosis screening of employees.

    Science.gov (United States)

    Kowada, Akiko

    2011-12-01

    Currently, an annual chest X-ray examination (CXR) for detection of active tuberculosis (TB) in employees aged ≥40 years is recommended in the guidelines of the Japan Industrial Safety and Health Law. Interferon-γ release assays are new alternatives to the tuberculin skin test for detecting Mycobacterium tuberculosis infection, with higher specificity than the tuberculin skin test and without cross-reactivity with the Bacille Calmette-Guérin vaccine. This study aimed to assess the cost-effectiveness of employee TB screening using QuantiFERON-TB Gold In-Tube (QFT) versus CXR. Markov models were constructed. The target population was a hypothetical cohort of immunocompetent 40-year-old individuals, using a societal perspective and a lifetime horizon. All costs and clinical benefits were discounted at a fixed annual rate of 3%. In a base-case analysis, the QFT strategy was the most cost-effective ($US 262.84; 22.87049 quality-adjusted life-years [QALYs]) compared with no screening ($448.38; 22.85452 QALYs) and CXR ($543.50; 22.85453 QALYs) [year 2009 values]. The QFT strategy is currently robust for screening Bacille Calmette-Guérin- vaccinated employees in Japan. There appears to be little role for CXR. These findings may be applicable to other countries in terms of choosing optimal TB screening for employees. Copyright © 2011 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Mosby, Inc. All rights reserved.

  3. Inhibition of clone formation as an assay for T cell-mediated cytotoxicity: short-term kinetics and comparison with 51Cr release

    International Nuclear Information System (INIS)

    Lees, R.K.; MacDonald, H.R.; Sinclair, N.R.; University of Western Ontario London

    1977-01-01

    The short-term kinetics of T cell-mediated cytotoxicity was investigated using a cloning inhibition assay. Murine cytotoxic thymus-derived lymphocytes generated in vitro in mixed leukocyte cultures were incubated for various periods of time at 37degC with allogeneic mastocytoma target cells. The mixtures were then plated in soft agar, and mastocytoma clone formation was assessed after 5-7 days incubation. Using this technique, it was demonstrated that events leading to the loss of cloning ability could be detected after 1-3 min incubation at 37degC, and after 20-30 min, 95% of the clone forming cells had been inactivated. When these results were compared directly with those obtained using the conventional 51 Cr-release assay, it was found that the events leading to loss of cloning ability occurred more rapidly than indicated by the isotope assay. However, a modification of the 51 Cr-release assay involving EDTA addition gave comparable result to the cloning inhibition assay. These results raise the possibility that the events leading to 51 Cr-release of tumor target cells may be related in time to those leading to the loss of cloning ability

  4. A High Throughput Screening Assay for Anti-Mycobacterial Small Molecules Based on Adenylate Kinase Release as a Reporter of Cell Lysis.

    Directory of Open Access Journals (Sweden)

    Lauren Forbes

    Full Text Available Mycobacterium tuberculosis (Mtb is well-established to be one of the most important bacterial pathogens for which new antimicrobial therapies are needed. Herein, we describe the development of a high throughput screening assay for the identification of molecules that are bactericidal against Mycobacteria. The assay utilizes the release of the intracellular enzyme adenylate kinase into the culture medium as a reporter of mycobacterial cell death. We demonstrate that the assay is selective for mycobactericidal molecules and detects anti-mycobacterial activity at concentrations below the minimum inhibitory concentration of many molecules. Thus, the AK assay is more sensitive than traditional growth assays. We have validated the AK assay in the HTS setting using the Mtb surrogate organism M. smegmatis and libraries of FDA approved drugs as well as a commercially available Diversity set. The screen of the FDA-approved library demonstrated that the AK assay is able to identify the vast majority of drugs with known mycobactericidal activity. Importantly, our screen of the Diversity set revealed that the increased sensitivity of the AK assay increases the ability of M. smegmatis-based screens to detect molecules with relatively poor activity against M. smegmatis but good to excellent activity against Mtb.

  5. Factors associated with the performance of a blood-based interferon-γ release assay in diagnosing tuberculosis.

    Directory of Open Access Journals (Sweden)

    Sally Banfield

    Full Text Available BACKGROUND: Indeterminate results are a recognised limitation of interferon-γ release assays (IGRA in the diagnosis of latent tuberculosis (TB infection (LTBI and TB disease, especially in children. We investigated whether age and common co-morbidities were associated with IGRA performance in an unselected cohort of resettled refugees. METHODS: A retrospective cross-sectional study of refugees presenting for their post-resettlement health assessment during 2006 and 2007. Refugees were investigated for prevalent infectious diseases, including TB, and for common nutritional deficiencies and haematological abnormalities as part of standard clinical screening protocols. Tuberculosis screening was performed by IGRA; QuantiFERON-TB Gold in 2006 and QuantiFERON-TBGold In-Tube in 2007. RESULTS: Complete data were available on 1130 refugees, of whom 573 (51% were children less than 17 years and 1041 (92% were from sub-Saharan Africa. All individuals were HIV negative. A definitive IGRA result was obtained in 1004 (89% refugees, 264 (26% of which were positive; 256 (97% had LTBI and 8 (3% had TB disease. An indeterminate IGRA result was obtained in 126 (11% refugees (all failed positive mitogen control. In multivariate analysis, younger age (linear OR= 0.93 [95% CI 0.91-0.95], P<0.001, iron deficiency anaemia (2.69 [1.51-4.80], P = 0.001, malaria infection (3.04 [1.51-6.09], P = 0.002, and helminth infection (2.26 [1.48-3.46], P<0.001, but not vitamin D deficiency or insufficiency, were associated with an indeterminate IGRA result. CONCLUSIONS: Younger age and a number of common co-morbidities are significantly and independently associated with indeterminate IGRA results in resettled predominantly African refugees.

  6. Evaluation of the tuberculin skin test and the interferon-γ release assay for TB screening in French healthcare workers

    Directory of Open Access Journals (Sweden)

    Raffi Francois

    2009-11-01

    Full Text Available Abstract Introduction Using French cut-offs for the Tuberculin Skin Test (TST, results of the TST were compared with the results of an Interferon-γ Release Assay (IGRA in Healthcare Workers (HCW after contact to AFB-positive TB patients. Methods Between May 2006 and May 2007, a total of 148 HCWs of the University Hospital in Nantes, France were tested simultaneously with IGRA und TST. A TST was considered to indicate recent latent TB infection (LTBI if an increase of >10 mm or if TST ≥ 15 mm for those with no previous TST result was observed. For those with a positive TST, chest X-ray was performed and preventive chemotherapy was offered. Results All HCWs were BCG-vaccinated. The IGRA was positive in 18.9% and TST ≥ 10 mm was observed in 65.5%. A recent LTBI was believed to be highly probable in 30.4% following TST. Agreement between IGRA and TST was low (kappa 0.041. In 10 (16.7% out of 60 HCWs who needed chest X-ray following TST the IGRA was positive. In 9 (20% out of 45 HCWs to whom preventive chemotherapy was offered following TST the IGRA was positive. Of those considered TST-negative following the French guidelines, 20.5% were IGRA-positive. In a two-step strategy - positive TST verified by IGRA - 18 out of 28 (64.3% IGRA-positive HCWs would not have been detected using French guidelines for TST interpretation. Conclusion The introduction of IGRA in contact tracings of BCG-vaccinated HCWs reduces X-rays and preventive chemotherapies. Increasing the cut-off for a positive TST does not seem to be helpful to overcome the effect of BCG vaccination on TST.

  7. The Combined Effects of Training on Serum Levels of Interferon Gamma (INF-γ and Expanded Scale Disability Status Scale of Patients with Multiple Sclerosis at Different Levels of Disability

    Directory of Open Access Journals (Sweden)

    Z Saberi

    2017-02-01

    Full Text Available Background and aim: Multiple sclerosis is a chronic and debilitating nervous system, leading to demyelination of the central nervous system (brain and spinal cord. Regular exercise and general physical activity is important to maintain health and prevent disease, already well known. Therefore the aim of this study was to evaluate the effect of 12 weeks of combined exercises (strength training, Strengthening Exercises, cardio respiratory endurance, a variety of static and dynamic balance exercises, exercises of the trunk (pilates training and walking on the treadmill training with body weight support on interferon gamma and Expanded Disability Status Scale women with multiple sclerosis. Methods: In the present experimental rsearch, female patients who were admitted to the MS Society of Shahrekord, Iran, were divided into three groups based on physical disability scores. In the first group (physical disability scale less than 4.5, 44 people were randomly selected to one experimental group (22 patients and control group (n = 22. In the second group (scale physical disability between 5 and 5.6, 26 patients were enrolled and randomly assigned to an experimental group (n = 13 and control group (n = 13. The third (Physical Disability Scale-up to 6.5, 26 patients were enrolled and randomly assigned to an experimental group (n = 13 and control group (n = 13. A total of 96 patients were participated in this study. Experimental groups of first, second and third were done its own intervention separately. While the control group received stretching exercises, workout schedule for the experimental group was of 12 weeks, three sessions of lasted one hour. Anthropometric factors and interferon-gamma were measured before and after training with the appropriate tools. Serum levels of INF-γ was determind using a commercial ELISA kit and EDSS scores were measured using the measure of disability in patients with MS. Data analysis was performed using descriptive

  8. Prevalence of scrub typhus in pyrexia of unknown origin and assessment of interleukin-8, tumor necrosis factor-alpha, and interferon-gamma levels in scrub typhus-positive patients.

    Science.gov (United States)

    Rizvi, Meher; Sultan, Asfia; Chowdhry, Madhav; Azam, Mohd; Khan, Fatima; Shukla, Indu; Khan, Haris M

    2018-01-01

    Scrub typhus is lesser known cause of fever of unknown origin in India. Even if there have been reports documenting the prevalence of scrub typhus in different parts of India, it is still an unknown entity, and clinicians usually do not consider it as differential diagnosis. The present study was performed to document the prevalence of scrub typhus among febrile patients in western part of Uttar Pradesh and to assess the clinical profile of infected patients on the one hand and knowledge, attitude, and practices among clinicians on the other. A total of 357 adult patients with fever of more than 5-day duration were recruited. All patients underwent complete physical examination, and detailed clinical history was elicited as per predesigned pro forma. After primary screening to rule out malaria, enteric fever, and leptospirosis infection, secondary screening for scrub typhus was done by rapid screen test and IgM ELISA. Scrub typhus infection was positive in 91 (25.5%) cases. The most common symptoms among the patients were fever (100%), pain in abdomen (79.1%), pedal edema 56 (61.5%), rash 44 (48.3%), headache 44 (48.3%), vomiting 42 (46.1%), constipation 33 (36.2%), cough 28 (30.7%), and lymphadenopathy 20 (21.9%). The median values of interleukin-8, interferon-gamma, and tumor necrosis factor-alpha in healthy controls were 15.54 pg/ml, 7.77 pg/ml, and 54.1 pg/ml, respectively, while the median values of these cytokines in scrub typhus-positive patients were 21.04 pg/ml, 8.74 pg/ml, and 73.8 pg/ml, respectively. Our results highlight that scrub typhus infection is an important cause of pyrexia of unknown origin, and active surveillance is necessary to assess the exact magnitude and distribution of the disease.

  9. Measurement of ex vivo ELISpot interferon-gamma recall responses to Plasmodium falciparum AMA1 and CSP in Ghanaian adults with natural exposure to malaria.

    Science.gov (United States)

    Ganeshan, Harini; Kusi, Kwadwo A; Anum, Dorothy; Hollingdale, Michael R; Peters, Bjoern; Kim, Yohan; Tetteh, John K A; Ofori, Michael F; Gyan, Ben A; Koram, Kwadwo A; Huang, Jun; Belmonte, Maria; Banania, Jo Glenna; Dodoo, Daniel; Villasante, Eileen; Sedegah, Martha

    2016-02-01

    Malaria eradication requires a concerted approach involving all available control tools, and an effective vaccine would complement these efforts. An effective malaria vaccine should be able to induce protective immune responses in a genetically diverse population. Identification of immunodominant T cell epitopes will assist in determining if candidate vaccines will be immunogenic in malaria-endemic areas. This study therefore investigated whether class I-restricted T cell epitopes of two leading malaria vaccine antigens, Plasmodium falciparum circumsporozoite protein (CSP) and apical membrane antigen-1 (AMA1), could recall T cell interferon-γ responses from naturally exposed subjects using ex vivo ELISpot assays. Thirty-five subjects aged between 24 and 43 years were recruited from a malaria-endemic urban community of Ghana in 2011, and their peripheral blood mononuclear cells (PBMCs) were tested in ELISpot IFN-γ assays against overlapping 15mer peptide pools spanning the entire CSP and AMA1 antigens, and 9-10mer peptide epitope mixtures that included previously identified and/or predicted human leukocyte antigen (HLA) class 1-restricted epitopes from same two antigens. For CSP, 26 % of subjects responded to at least one of the nine 15mer peptide pools whilst 17 % responded to at least one of the five 9-10mer HLA-restricted epitope mixtures. For AMA1, 63 % of subjects responded to at least one of the 12 AMA1 15mer peptide pools and 51 % responded to at least one of the six 9-10mer HLA-restricted epitope mixtures. Following analysis of data from the two sets of peptide pools, along with bioinformatics predictions of class I-restricted epitopes and the HLA supertypes expressed by a subset of study subjects, peptide pools that may contain epitopes recognized by multiple HLA supertypes were identified. Collectively, these results suggest that natural transmission elicits ELISpot IFN-γ activities to class 1-restricted epitopes that are largely HLA-promiscuous. These

  10. High-sensitive and rapid detection of Mycobacterium tuberculosis infection by IFN-γ release assay among HIV-infected individuals in BCG-vaccinated area

    Directory of Open Access Journals (Sweden)

    Jiang Weimin

    2009-05-01

    Full Text Available Abstract Background An accurate test for Mycobacterium tuberculosis infection is urgently needed in immunosuppressed populations. The aim of this study was to investigate the diagnostic power of enzyme-linked immunospot (ELISPOT-based IFN-γ release assay in detecting active and latent tuberculosis in HIV-infected population in bacillus Calmette-Guerin (BCG-vaccinated area. A total of 100 HIV-infected individuals including 32 active tuberculosis patients were recruited. An ELISPOT-based IFN-γ release assay, T-SPOT.TB, was used to evaluate the M. tuberculosis ESAT-6 and CFP-10 specific IFN-γ response. Tuberculin skin test (TST was performed for all recruited subjects. Results The subjects were divided into group HIV+ATB (HIV-infected individuals with active tuberculosis, n = 32, group HIV+LTB (HIV-infected individuals with positive results of T-SPOT.TB assay, n = 46 and group HIV only (HIV-infected individuals with negative results of T-SPOT.TB assay and without evidence of tuberculosis infection, n = 22. In group HIV+ATB and HIV+LTB, T-SPOT.TB positive rate in subjects with TST P 85% in patients with TB treatment for less than 1 month and CD4+ T cells ≥200/μl, while for patients treated for more than 3 months and CD4+ T cells Conclusion ELISPOT-based IFN-γ release assay is more sensitive and rapid for the diagnosis of TB infection in Chinese HIV-infected individuals with history of BCG vaccination, and could be an effective tool for guiding preventive treatment with isoniazid in latently infected people and for TB control in China.

  11. [CLINICAL UTILITY OF T-SPOT.TB ASSAY WITH T-Cell Xtend REAGENT FOR ACTIVE TUBERCULOSIS DIAGNOSIS IN THE FIELD TEST AT OUR HOSPITAL].

    Science.gov (United States)

    Nemoto, Kenji; Oh-ishi, Shuji; Taguchi, Masato; Hyodo, Kentaro; Kanazawa, Jun; Miura, Yukiko; Takaku, Takio; Usui, Shingo; Hayashihara, Kenji; Saito, Takefumi

    2016-04-01

    T-SPOT.TB (T-SPOT), an interferon-gamma release assay, has shown promise as a diagnostic tool for active tuberculosis (TB), and its use is expanding. Addition of the T-Cell Xtend (TCX) reagent may allow delayed processing, and this characteristic is important for using this test in the field. However, limited data is available on the usefulness of T-SPOT with TCX as a field test for diagnosing active TB. To investigate the clinical utility of T-SPOT with TCX and the risk factors for a false-negative result in patients with active TB. A total of 57 patients with active TB who underwent the T-SPOT test with TCX prior to treatment were enrolled between May 2013 and May 2015. One patient with an indeterminate result for T-SPOT was excluded; therefore, the data of 56 patients were eventually included in the final analysis. The basic characteristics and clinical findings were compared between the true-positive and false-negative T-SPOT groups. Of the 56 patients, 40 (71.4%), 13 (23.2%), 3 (5.4%) had true-positive, false-negative, and borderline T-SPOT results, respectively. This study did not reveal any significant risk factors for a false-negative T-SPOT result. In this clinical study, the proportion of patients with a false-negative result for T-SPOT with TCX for active TB was higher than that reported previously. Therefore, careful interpretation of a negative result for T-SPOT with TCX is necessary, regardless of the patient's background.

  12. A new assay for cytotoxic lymphocytes, based on a radioautographic readout of 111In release, suitable for rapid, semi-automated assessment of limit-dilution cultures

    International Nuclear Information System (INIS)

    Shortman, K.; Wilson, A.

    1981-01-01

    A new assay for cytotoxic T lymphocytes is described, of general application, but particularly suitable for rapid, semi-automated assessment of multiple microculture tests. Target cells are labelled with high efficiency and to high specific activity with the oxine chelate of 111 indium. After a 3-4 h incubation of test cells with 5 X 10 3 labelled target cells in V wells of microtitre trays, samples of the supernatant are spotted on paper (5 μl) or transferred to soft-plastic U wells (25-50 μl) and the 111 In release assessed by radioautography. Overnight exposure of X-ray film with intensifying screens at -70 0 C gives an image which is an intense dark spot for maximum release, a barely visible darkening with the low spontaneous release, and a definite positive with 10% specific lysis. The degree of film darkening, which can be quantitated by microdensitometry, shows a linear relationship with cytotoxic T lymphocyte dose up to the 40% lysis level. The labelling intensity and sensitivity can be adjusted over a wide range, allowing a single batch of the short half-life isotope to serve for 2 weeks. The 96 assays from a single tray are developed simultaneously on a single small sheet of film. Many trays can be processed together, and handling is rapid if 96-channel automatic pipettors are used. The method allows rapid visual scanning for positive and negative limit dilution cultures in cytotoxic T cell precursor frequency and specificity studies. In addition, in conjunction with an automated densitometer designed to scan microtitre trays, the method provides an efficient alternative to isotope counting in routine cytotoxic assays. (Auth.)

  13. A method of radiocompetitive assay of total thyroxine in the serum by means of enzymatic release of thyroxine from the transporting proteins

    International Nuclear Information System (INIS)

    Snarski, A.; Wyrwinski, J.

    1978-01-01

    Pepsin causes denaturation of the transporting proteins and liberates thyroxine which can be assayed by the radiocompetitive method. Change of the pH of the medium from acid to alkaline inactivates irreveribly pepsin. The enzymatic release of thyroxine is much simpler that the method of ethanol extraction and thermal denaturation of the transporting proteins applied up to now. The new technique of thyroxine release has been introduced for radiocompetitive determination of thyroxine using dextran coated charcoal for adsorption of the free hormone. A new method has been elaborated for preparation of working standards of thyroxine in a mixture of pepsin solution with hormone-free serum. The method is efficient and rapid. The normal range is from 50 to 130 nanomol/l. Over 7 000 determinations were done as yet in patients with suspected thyroid function disturbances. (author)

  14. Interferon-γ release assays for the diagnosis of latent Mycobacterium tuberculosis infection: a systematic review and meta-analysis

    DEFF Research Database (Denmark)

    Diel, R; Goletti, D; Ferrara, G

    2011-01-01

    We conducted a systematic review and meta-analysis to compare the accuracy of the QuantiFERON-TB® Gold In-Tube (QFT-G-IT) and the T-SPOT®.TB assays with the tuberculin skin test (TST) for the diagnosis of latent Mycobacterium tuberculosis infection (LTBI). The Medline, Embase and Cochrane databases...... of IGRAs varied 98-100%. In immunocompetent adults, NPV for progression to tuberculosis within 2 yrs were 97.8% for T-SPOT®.TB and 99.8% for QFT-G-IT. When test performance of an immunodiagnostic test was not restricted to prior positivity of another test, progression rates to tuberculosis among IGRA...

  15. Cerebrospinal fluid interferon-gamma in the diagnosis of tuberculous meningitis%脑脊液γ-干扰素检测在结核性脑膜炎中的临床价值

    Institute of Scientific and Technical Information of China (English)

    黄锡坤; 欧结艳

    2014-01-01

    目的:探讨血清及脑脊液的γ-干扰素(I N F-γ)水平对结核性脑膜炎诊断的临床价值。方法采用酶联免疫吸附试验(ELISA)检测25例结核性脑膜炎(结核性脑膜炎组)、31例病毒性脑膜炎(病毒性脑膜炎组)、12例化脓性脑膜炎(化脓性脑膜炎组)患者血清及脑脊液中IN F-γ水平。结果结核性脑膜炎组患者脑脊液IN F-γ含量为(386.3±83.5)ng/L ,明显高于病毒性脑膜炎组(63.4±16.2)ng/L和化脓性脑膜炎组(116.8±22.6)ng/L ,差异有统计学意义( P<0.01),且重叠性很小;而3组血清INF-γ含量很低并相近,差异无统计学意义(P>0.05)。脑脊液INF-γ诊断结核性脑膜炎的灵敏度、特异度和准确度分别为92.0%、95.3%和94.1%。结论检测脑脊液IN F-γ对结核性脑膜炎有一定的辅助诊断价值。%Objective To evaluate the clinical utility of interferon-gamma (IFN-γ) in serum and cerebrospinal fluid in the diagnosis of tuberculous meningitis .Methods IFN-γlevels in serum and cerebrospinal fluid were analyzed by ELISA method in 25 patients with tuberculous meningitis ,31 patients with viral meningitis and 12 patients with suppurative meningitis .Results The IFN-γlevels in cerebrospinal fluid were significantly higher in the patients with tuberculous meningitis (386 .3 ± 83 .5) ng/L than in the patients with viral meningitis (63 .4 ± 16 .2) ng/L or the patients with suppurative meningitis (116 .8 ± 22 .6) ng/L (P0 .05) .The sensitivity ,specificity and accuracy of IFN-γ level in cerebrospinal fluid for diagnosis of tuberculous meningitis were 92 .0% ,95 .3% and 94 .1% ,respectively .Conclusions Measurement of IFN-γlevel in cerebrospinal fluid may be helpful for the diagnosis of tuberculous meningitis .

  16. Revisiting the IFN-γ release assay: Whole blood or PBMC cultures? - And other factors of influence

    DEFF Research Database (Denmark)

    Hartmann, Sofie Bruun; Emnéus, Jenny; Wolff, Anders

    2016-01-01

    light on external factors that could influence the read out in terms of IFN-γ levels. It was found that optimal culture conditions varied between individual animals; when polyclonal activated, cells from whole blood cultures were most responsive, but when activated specifically, the optimal cell....... However, there is no consensus whether to use whole blood cultures or purified PBMCs for the assay, and both cell populations are being used and results compared. Therefore the aim of this study was to compare different culture settings using immune cells from previously vaccinated calves, and to shed...... concentration/population varied with whole blood, 10 × 106 cells/ml PBMC and 5 × 106 cells/ml PBMC being the highest performing conditions. A further investigation of the distribution of cell populations in PBMCs compared to whole blood was conducted, and a significant (p

  17. A survey of monitoring and assay systems for release of metals from radiation controlled areas at LANL.

    Energy Technology Data Exchange (ETDEWEB)

    Gruetzmacher, K. M. (Kathleen M.); MacArthur, D. W. (Duncan W.)

    2002-01-01

    At Los Alamos National Laboratory (LANL), a recent effort in waste minimization has focused on scrap metal from radiological controlled areas (RCAs). In particular, scrap metal from RCAs needs to be dispositioned in a reasonable and cost effective manner. Recycling of DOE scrap metals from RCAs is currently under a self-imposed moratorium. Since recycling is not available and reuse is difficult, often metal waste from RCAs, which could otherwise be recycled, is disposed of as low-level waste. Estimates at LANL put the cost of low-level waste disposal at $550 to $4000 per cubic meter, depending on the type of waste and the disposal site. If the waste is mixed, the cost for treatment and disposal can be as high as $50,000 per cubic meter. Disposal of scrap metal as low-level waste uses up valuable space in the low-level waste disposal areas and requires transportation to the disposal site under Department of Transportation (DOT) regulations for low-level waste. In contrast, disposal as non-radioactive waste costs as little as $2 per cubic meter. While recycling is unavailable, disposing of the metal at an industrial waste site could be the best solution for this waste stream. A Green Is Clean (GIC) type verification program needs to be in place to provide the greatest assurance that the waste does not contain DOE added radioactivity. This paper is a review of available and emerging radiation monitoring and assay systems that could be used for scrap metal as part of the LANL GIC program.

  18. Screening health care workers with interferon-γ release assay versus tuberculin skin test: impact on costs and adherence to testing (the SWITCH study).

    Science.gov (United States)

    Wrighton-Smith, Peter; Sneed, Laurie; Humphrey, Frances; Tao, Xuguang; Bernacki, Edward

    2012-07-01

    To determine the price point at which an interferon-γ release assay (IGRA) is less costly than a tuberculin skin test (TST) for health care employee tuberculosis screening. A multidecision tree-based cost model incorporating inputs gathered from time-motion studies and parallel testing by IGRA and TST was conducted in a subset of our employees. Administering a TST testing program costs $73.20 per person screened, $90.80 per new hire, and $63.42 per annual screen. Use of an IGRA for employee health testing is cost saving at an IGRA test cost of $54.83 or less per test and resulted in higher completion rates because of the elimination of the need for a second visit to interpret the TST. Using an IGRA for employee health screening can be an institutional cost saving and results in higher compliance rates.

  19. IL-8、 MMP-9、 INF-γ的检测对结核性脑膜炎及病毒性脑膜炎发病的意义%Detection of interleukin-8, matrix metalloproteinase-9 and interferon gamma levels in the cerebrospinal fluid of patients with tuberculous meningitis and viral meningitis

    Institute of Scientific and Technical Information of China (English)

    朱飞; 张家堂; 邢小微; 贺路星; 赵威; 郎森阳; 于生元

    2012-01-01

    目的 探讨脑脊液中白细胞介素-8(IL-8)、基质金属蛋白酶-9(MMP-9)、干扰素-γ(INF-γ)含量的检测对结核性脑膜炎及病毒性脑膜炎的临床诊断价值. 方法 选取解放军总医院、解放军第三0九医院自2010年8月至2011年11月住院的患者,其中结核性脑膜炎组20例,病毒性脑膜炎组15例,非感染性神经系统疾病组20例.用ELISA法检测3组患者脑脊液IL-8、MMP-9、INF-γ含量,并进行比较分析. 结果 结核性脑膜炎组患者脑脊液中IL-8、MMP-9、INF-γ的含量高于病毒性脑膜炎组和非感染性神经系统疾病组差异有统计学意义(P<0.05).病毒性脑膜炎组患者脑脊液中IL-8、MMP-9含量高于非感染性神经系统组(P<0.05).病毒性脑膜炎组患者脑脊液中INF-γ含量与非感染性神经系统疾病组比较差异无统计学意义(P>0.05). 结论 脑脊液中IL-8、MMP-9、INF-γ含量的检测对结核性脑膜炎具有一定的辅助诊断意义.IL-8、MMP-9在病毒性脑膜炎的发病和进展中亦起到一定作用.临床上若在患者脑脊液中检测到高水平的INF-γ,较之IL-8、MMP-9对于结核性脑膜炎更具诊断价值.%Objective To investigate the diagnostic values of interleukin-8 (IL-8), matrix metalloproteinase-9 (MMP-9) and interferon gamma (INF-γ) levels in patients with tuberculous meningitis and viral meningitis by detecting the contents of these biomarkers in the cerebrospinal fluid (CSF). Methods Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of IL-8,MMP-9 and INF-γ in the CSF of patients with tuberculous meningitis (n=20),viral meningitis (n=15) and noninfectious neurologic diseases (n=20) who admitted to our hospital from August 2010 to November 2011. Results The IL-8,MMP-9 and INF-γlevels in the samples from the tuberculous meningitis patients were significantly higher than those from either viral meningitis or noninfectious neurologic diseases (P<0.05).The contents of IL-8

  20. Assay of 6-gingerol in CO2 supercritical fluid extracts of ginger and evaluation of its sustained release from a transdermal delivery system across rat skin.

    Science.gov (United States)

    Chen, Yan; Zhang, Cuiping; Zhang, Mei; Fu, Xiaobing

    2014-07-01

    Ginger has been widely used as healthy food condiment as well as traditional Chinese medicine since antiquity. Multiple potentials of ginger for treatment of various ailments have been revealed. However, the biological half-life of 6-gingerol (a principal pungent ingredient of ginger) is only 7.23 minutes while taken orally. Delivery of ginger compositions by routes other than oral have scarcely been reported. Therefore, we studied a noninvasive transdermal drug delivery system (TDDS) of ginger to bypass hepatic first pass metabolism, avoid gastrointestinal degradation and achieve long persistent release of effective compositions. After establishment of a HPLC analysis method of 6-gingerol, assays of 6-gingerol were performed to compare two kinds of ginger extracts. Then, the characteristics of transdermal delivery of 6-gingerol in TDDS were exhibited. The results showed that the contents of 6-gingerol in two kinds of ginger extracts were significantly different. The maximal delivery percentage of 6-gingerol across rat skin at 20 h was more than 40% in different TDDS formulations. TDDS may provide long-lasting delivery of ginger compounds.

  1. Whole blood assay to access T cell-immune responses to Mycobacterium tuberculosis antigens in healthy Brazilian individuals

    Directory of Open Access Journals (Sweden)

    Paulo RZ Antas

    2004-02-01

    Full Text Available The production of interferon gamma (IFNgamma guarantees effective T cell-mediated immunity against Mycobacterium tuberculosis infection. In the present study, we simply compare the in vitro immune responses to Mycobacterium antigens in terms of IFNg production in a total of 10 healthy Brazilian volunteers. Whole blood and mononuclear cells were cultivated in parallel with PPD, Ag85B, and M. bovis hsp65, and five-days supernatants were harvested for cytokine detection by ELISA. The inter-assay result was that the overall profile of agreement in response to antigens was highly correlated (r² = 0.9266; p = 0.0102. Potential analysis is in current progress to dictate the usefulness of this method to access the immune responses also in tuberculosis patients and its contacts.

  2. Hormone assay

    International Nuclear Information System (INIS)

    Eisentraut, A.M.

    1977-01-01

    An improved radioimmunoassay is described for measuring total triiodothyronine or total thyroxine levels in a sample of serum containing free endogenous thyroid hormone and endogenous thyroid hormone bound to thyroid hormone binding protein. The thyroid hormone is released from the protein by adding hydrochloric acid to the serum. The pH of the separated thyroid hormone and thyroid hormone binding protein is raised in the absence of a blocking agent without interference from the endogenous protein. 125 I-labelled thyroid hormone and thyroid hormone antibodies are added to the mixture, allowing the labelled and unlabelled thyroid hormone and the thyroid hormone antibody to bind competitively. This results in free thyroid hormone being separated from antibody bound thyroid hormone and thus the unknown quantity of thyroid hormone may be determined. A thyroid hormone test assay kit is described for this radioimmunoassay. It provides a 'single tube' assay which does not require blocking agents for endogenous protein interference nor an external solid phase sorption step for the separation of bound and free hormone after the competitive binding step; it also requires a minimum number of manipulative steps. Examples of the assay are given to illustrate the reproducibility, linearity and specificity of the assay. (UK)

  3. Comparison of interferon-γ release assay to two cut-off points of tuberculin skin test to detect latent Mycobacterium tuberculosis infection in primary health care workers.

    Directory of Open Access Journals (Sweden)

    Fernanda Mattos de Souza

    Full Text Available An interferon-γ release assay, QuantiFERON-TB (QFT test, has been introduced an alternative test for the diagnosis of latent Mycobacterium tuberculosis infection (LTBI. Here, we compared the performance of QFT with tuberculin skin test (TST measured at two different cut-off points among primary health care work (HCW in Brazil.A cross-sectional study was carried out among HCWs in four Brazilian cities with a known history of high incidence of TB. Results of the QFT were compared to TST results based on both ≥5 mm and ≥10 mm as cut-off points.We enrolled 632 HCWs. When the cut-off value of ≥10 mm was used, agreement between QFT and TST was 69% (k = 0.31, and when the cut-off of ≥5 mm was chosen, the agreement was 57% (k = 0.22. We investigated possible factors of discordance of TST vs QFT. Compared to the TST-/QFT- group, risk factors for discordance in the TST+/QFT- group with TST cut-off of ≥5 mm included age between 41-45 years [OR = 2.70; CI 95%: 1.32-5.51] and 46-64 years [OR = 2.04; CI 95%: 1.05-3.93], BCG scar [OR = 2.72; CI 95%: 1.40-5.25], and having worked only in primary health care [OR = 2.30; CI 95%: 1.09-4.86]. On the other hand, for the cut-off of ≥10 mm, BCG scar [OR = 2.26; CI 95%: 1.03-4.91], being a household contact of a TB patient [OR = 1.72; CI 95%: 1.01-2.92] and having had a previous TST [OR = 1.66; CI 95%: 1.05-2.62], were significantly associated with the TST+/QFT- group. No statistically significant associations were found among the TST-/QFT+ discordant group with either TST cut-off value.Although we identified BCG vaccination to contribute to the discordance at both TST cut-off measures, the current Brazilian recommendation for the initiation of LTBI treatment, based on information gathered from medical history, TST, chest radiograph and physical examination, should not be changed.

  4. Comparison of interferon-γ release assay to two cut-off points of tuberculin skin test to detect latent Mycobacterium tuberculosis infection in primary health care workers.

    Science.gov (United States)

    de Souza, Fernanda Mattos; do Prado, Thiago Nascimento; Pinheiro, Jair dos Santos; Peres, Renata Lyrio; Lacerda, Thamy Carvalho; Loureiro, Rafaela Borge; Carvalho, Jose Américo; Fregona, Geisa; Dias, Elias Santos; Cosme, Lorrayne Beliqui; Rodrigues, Rodrigo Ribeiro; Riley, Lee Wood; Maciel, Ethel Leonor Noia

    2014-01-01

    An interferon-γ release assay, QuantiFERON-TB (QFT) test, has been introduced an alternative test for the diagnosis of latent Mycobacterium tuberculosis infection (LTBI). Here, we compared the performance of QFT with tuberculin skin test (TST) measured at two different cut-off points among primary health care work (HCW) in Brazil. A cross-sectional study was carried out among HCWs in four Brazilian cities with a known history of high incidence of TB. Results of the QFT were compared to TST results based on both ≥5 mm and ≥10 mm as cut-off points. We enrolled 632 HCWs. When the cut-off value of ≥10 mm was used, agreement between QFT and TST was 69% (k = 0.31), and when the cut-off of ≥5 mm was chosen, the agreement was 57% (k = 0.22). We investigated possible factors of discordance of TST vs QFT. Compared to the TST-/QFT- group, risk factors for discordance in the TST+/QFT- group with TST cut-off of ≥5 mm included age between 41-45 years [OR = 2.70; CI 95%: 1.32-5.51] and 46-64 years [OR = 2.04; CI 95%: 1.05-3.93], BCG scar [OR = 2.72; CI 95%: 1.40-5.25], and having worked only in primary health care [OR = 2.30; CI 95%: 1.09-4.86]. On the other hand, for the cut-off of ≥10 mm, BCG scar [OR = 2.26; CI 95%: 1.03-4.91], being a household contact of a TB patient [OR = 1.72; CI 95%: 1.01-2.92] and having had a previous TST [OR = 1.66; CI 95%: 1.05-2.62], were significantly associated with the TST+/QFT- group. No statistically significant associations were found among the TST-/QFT+ discordant group with either TST cut-off value. Although we identified BCG vaccination to contribute to the discordance at both TST cut-off measures, the current Brazilian recommendation for the initiation of LTBI treatment, based on information gathered from medical history, TST, chest radiograph and physical examination, should not be changed.

  5. Tuberculosis Screening on a Health Science Campus: Use of QuantiFERON-TB Gold Test for Students and Employees

    Science.gov (United States)

    Veeser, Peggy Ingram; Smith, Phillip Karl; Handy, Barry; Martin, Sharon R.

    2007-01-01

    Detecting and managing "Mycobacterium tuberculosis" (TB) infection in a health-science center population is a clinical dilemma. Tuberculin skin tests are still the preferred method for detecting present or past infection of TB. The authors discuss the performance of whole blood interferon gamma release assay test commercially known as…

  6. Release patterns of pregnancy-associated plasma protein A in patients with acute coronary syndromes assessed by an optimized monoclonal antibody assay

    DEFF Research Database (Denmark)

    Schoos, Mikkel; Iversen, Kasper; Teisner, Ane

    2008-01-01

    Objective. Pregnancy-associated plasma protein A (PAPP-A) is expressed in eroded and ruptured atheromatous plaques, and circulating levels are elevated in acute coronary syndromes (ACS). Our objective was to investigate release patterns of PAPP-A in ACS and whether they differ among different typ...

  7. Interferon-gamma regulates oxidative stress during experimental autoimmune encephalomyelitis

    DEFF Research Database (Denmark)

    Espejo, Carmen; Penkowa, Milena; Sáez-Torres, Irene

    2002-01-01

    disease eliciting secretion of proinflammatory cytokines like IFN-gamma or TNF-alpha, and it has been suggested that cytokine-induced oxidative stress could have a role in EAE neuropathology. However, the individual roles of these and other cytokines in the pathogenesis of the disease are still uncertain....... Here we analyze the role of IFN-gamma during EAE by using both IFN-gamma receptor-knockout (IFN-gamma R(-/-)) and wild-type mice, both strains immunized with peptide 40-55 from rat myelin oligodendrocyte glycoprotein. The levels of oxidative stress were determined through the analysis...... of immunoreactivity for inducible NO synthase, nitrotyrosine, and malondialdehyde, as well as through the expression of the tissue-protective antioxidant factors metallothionein I+II (MT-I+II). We also examined the number of cells undergoing apoptosis as judged by using the TUNEL technique. The levels of oxidative...

  8. Expression of human interferon gamma in Brassica napus seeds

    African Journals Online (AJOL)

    TUOYO

    2010-08-09

    Aug 9, 2010 ... resulted band was purified using the agarose gel DNA extraction kit. (Roche). ..... rape seed napin structure and potential roles of the storage protein. ... the sensitivity of progressive multiple sequence alignment through.

  9. Interferon-gamma treatment kinetics among patients with active ...

    African Journals Online (AJOL)

    Introduction: Interferon-γ (IFN-γ) is essential for defence against Mycobacterium tuberculosis; however, levels in patients with active tuberculosis (TB) and changes during treatment have not been documented in our tuberculosis patients in Nigeria, hence this study has been carried out. Objective: To determine variations, ...

  10. A strategy for the unrestricted release of metallic scrap from decommissioned nuclear facilities, integrating quality planning with the effective use of non-destructive assay instrumentation

    International Nuclear Information System (INIS)

    Gunn, R.; Troughten, N.; Warran, C.

    2000-01-01

    The key component of any release operation is its quality plan. This plan is a step by step procedure which describes all the actions which are required during the entire process and also the information required at every stage of the process. Once drafted the quality plan will enable operators to determine at what stages it is necessary to measure the parameters required to allow the process to progress. The objective of the radiometric system is to provide the auditable proof that material has been surveyed, the record of the survey and the evidence that the system was operating within its design parameters during the survey. Typical radiometric instruments which have been deployed to provide the information requirements of the quality plan are described in this paper. These include the IonSens TM Conveyor, conveyorized survey monitor used to provide high throughout clearance surveys of removed metallic scrap. The IonSens TM Pipe, cut pipe monitor is used to survey interiors and exteriors of long lengths of removed pipe-work. The IonSens TM 208 large Item monitor is used to survey removed pieces of vessels and ductwork which may have convoluted shapes and finally the DrumScan TM system is used to survey 200 litre drums containing cut up pieces of metal. (author)

  11. A radioreceptor assay of luteinizing hormone-releasing hormone receptor and characterization of LHRH binding to pituitary receptors in Shao duck

    International Nuclear Information System (INIS)

    Yang Peixin; Wu Meiwen; Chen Ziyuan

    2000-01-01

    The properties of Shao duck pituitary luteinizing hormone-releasing hormone (LHRH) receptors were analyzed in pituitary membrane preparation and isolated pituitary cells prepared by enzymatic dispersion with collagenase and trypsin, by using a super-agonist analog of (D-Lys 6 ) LHRH. High binding of 125 I-(D-Lys 6 ) LHRH to 10 6 cultured cells of Shao duck was observed after a 90 minute incubation at 4 degree C, while binding was significantly reduced after a 24h incubation. Binding of the radioligand was a function of tissue concentration of Shao duck pituitary membrane preparation, with a positive correlation over the range of 1-2 pituitary per-tube. Specific binding for 125 I-(D-Lys 6 ) LHRH increased with the increase in the amount of 125 I-(D-Lys 6 ) LHRH. The Scatchard analysis of data revealed a linear relationship between the amount of specific binding and the ratio of specific binding to free 1 '2 5 I(D-Lys 6 )LHRH, indicating a single class of high affinity sites. Equilibrium dissociation constant (Kd) was 0.34 nM in pituitary membrane preparation and 0.43 nM in isolated pituitary cells. Both Kd values were near and the maximum binding capacity (B max ) was great in isolated cells, suggesting no significant loss of the LHRH receptor population caused by the enzymatic procedure employed for cell dispersion in the present study. Addition of 9D-Lys 6 ) LHRH displaced bound 125 I-(D-Lys 6 ) LHRH. These results demonstrated the presence and provided characterization of LHRH receptors in Shao duck pituitary

  12. Differential expression of interferon-gamma and interferon-gamma-inducing cytokines in Thai patients with scrub typhus or leptospirosis

    NARCIS (Netherlands)

    Chierakul, Wirongrong; de Fost, Maaike; Suputtamongkol, Yupin; Limpaiboon, Roongreung; Dondorp, Arjen; White, Nicholas J.; van der Poll, Tom

    2004-01-01

    Interferon (IFN)-gamma plays an important role in the induction of a type 1 immune response against intracellular pathogens. We compared the plasma levels of IFN-gamma and IFN-gamma-inducing cytokines in adult Thai patients with scrub typhus, caused by the obligate intracellular bacterium Orientia

  13. Mechanism and kinetics of the loss of poorly soluble drugs from liposomal carriers studied by a novel flow field-flow fractionation-based drug release-/transfer-assay.

    Science.gov (United States)

    Hinna, Askell Hvid; Hupfeld, Stefan; Kuntsche, Judith; Bauer-Brandl, Annette; Brandl, Martin

    2016-06-28

    Liposomes represent a versatile drug formulation approach e.g. for improving the water-solubility of poorly soluble drugs but also to achieve drug targeting and controlled release. For the latter applications it is essential that the drug remains associated with the liposomal carrier during transit in the vascular bed. A range of in vitro test methods has been suggested over the years for prediction of the release of drug from liposomal carriers. The majority of these fail to give a realistic prediction for poorly water-soluble drugs due to the intrinsic tendency of such compounds to remain associated with liposome bilayers even upon extensive dilution. Upon i.v. injection, in contrast, rapid drug loss often occurs due to drug transfer from the liposomal carriers to endogenous lipophilic sinks such as lipoproteins, plasma proteins or membranes of red blood cells and endothelial cells. Here we report on the application of a recently introduced in vitro predictive drug transfer assay based on incubation of the liposomal drug carrier with large multilamellar liposomes, the latter serving as a biomimetic model sink, using flow field-flow fractionation as a tool to separate the two types of liposomes. By quantifying the amount of drug remaining associated with the liposomal drug carrier as well as that transferred to the acceptor liposomes at distinct times of incubation, both the kinetics of drug transfer and release to the water phase could be established for the model drug p-THPP (5,10,15,20-tetrakis(4-hydroxyphenyl)21H,23H-porphine). p-THPP is structurally similar to temoporfin, a photosensitizer which is under clinical evaluation in a liposomal formulation. Mechanistic insights were gained by varying the donor-to-acceptor lipid mass ratio, size and lamellarity of the liposomes. Drug transfer kinetics from one liposome to another was found rate determining as compared to redistribution from the outermost to the inner concentric bilayers, such that the overall

  14. The risk of tuberculosis related to tumour necrosis factor antagonist therapies: a TBNET consensus statement

    DEFF Research Database (Denmark)

    Solovic, I.; Sester, M.; Gomez-Reino, J.J.

    2010-01-01

    risk of reactivating latent infections, especially tuberculosis (TB). Following TNF antagonist therapy, the relative risk for TB is increased up to 25 times, depending on the clinical setting and the TNF antagonist used. Interferon-gamma release assays or, as an alternative in individuals without...... a history of bacille Calmette-Guerin vaccination, tuberculin skin testing is recommended to screen all adult candidates for TNF antagonist treatment for the presence of latent infection with Mycobacterium tuberculosis. Moreover, paediatric practice suggests concomitant use of both the tuberculin skin test...... and an interferon-gamma release assay, as there are insufficient data in children to recommend one test over the other. Consequently, targeted preventive chemotherapy is highly recommended for all individuals with persistent M. tuberculosis-specific immune responses undergoing TNF antagonist therapy...

  15. Cross-validated stable-isotope dilution GC-MS and LC-MS/MS assays for monoacylglycerol lipase (MAGL) activity by measuring arachidonic acid released from the endocannabinoid 2-arachidonoyl glycerol.

    Science.gov (United States)

    Kayacelebi, Arslan Arinc; Schauerte, Celina; Kling, Katharina; Herbers, Jan; Beckmann, Bibiana; Engeli, Stefan; Jordan, Jens; Zoerner, Alexander A; Tsikas, Dimitrios

    2017-03-15

    2-Arachidonoyl glycerol (2AG) is an endocannabinoid that activates cannabinoid (CB) receptors CB1 and CB2. Monoacylglycerol lipase (MAGL) inactivates 2AG through hydrolysis to arachidonic acid (AA) and glycerol, thus modulating the activity at CB receptors. In the brain, AA released from 2AG by the action of MAGL serves as a substrate for cyclooxygenases which produce pro-inflammatory prostaglandins. Here we report stable-isotope GC-MS and LC-MS/MS assays for the reliable measurement of MAGL activity. The assays utilize deuterium-labeled 2AG (d 8 -2AG; 10μM) as the MAGL substrate and measure deuterium-labeled AA (d 8 -AA; range 0-1μM) as the MAGL product. Unlabelled AA (d 0 -AA, 1μM) serves as the internal standard. d 8 -AA and d 0 -AA are extracted from the aqueous buffered incubation mixtures by ethyl acetate. Upon solvent evaporation the residue is reconstituted in the mobile phase prior to LC-MS/MS analysis or in anhydrous acetonitrile for GC-MS analysis. LC-MS/MS analysis is performed in the negative electrospray ionization mode by selected-reaction monitoring the mass transitions [M-H] - →[M-H - CO 2 ] - , i.e., m/z 311→m/z 267 for d 8 -AA and m/z 303→m/z 259 for d 0 -AA. Prior to GC-MS analysis d 8 -AA and d 0 -AA were converted to their pentafluorobenzyl (PFB) esters by means of PFB-Br. GC-MS analysis is performed in the electron-capture negative-ion chemical ionization mode by selected-ion monitoring the ions [M-PFB] - , i.e., m/z 311 for d 8 -AA and m/z 303 for d 0 -AA. The GC-MS and LC-MS/MS assays were cross-validated. Linear regression analysis between the concentration (range, 0-1μM) of d 8 -AA measured by LC-MS/MS (y) and that by GC-MS (x) revealed a straight line (r 2 =0.9848) with the regression equation y=0.003+0.898x, indicating a good agreement. In dog liver, we detected MAGL activity that was inhibitable by the MAGL inhibitor JZL-184. Exogenous eicosatetraynoic acid is suitable as internal standard for the quantitative determination

  16. Assay system

    International Nuclear Information System (INIS)

    Patzke, J.B.; Rosenberg, B.J.

    1984-01-01

    The accuracy of assays for monitoring concentrations of basic drugs in biological fluids containing a 1 -acid glycoproteins, such as blood (serum or plasma), is improved by the addition of certain organic phosphate compounds to minimize the ''protein effect.'' Kits containing the elements of the invention are also disclosed

  17. Safety and efficacy of the C-Tb skin test to diagnose Mycobacterium tuberculosis infection, compared with an interferon γ release assay and the tuberculin skin test: a phase 3, double-blind, randomised, controlled trial.

    Science.gov (United States)

    Ruhwald, Morten; Aggerbeck, Henrik; Gallardo, Rafael Vázquez; Hoff, Søren T; Villate, José I; Borregaard, Bettine; Martinez, José A; Kromann, Ingrid; Penas, Antón; Anibarro, Luis L; de Souza-Galvão, Maria Luiza; Sánchez, Francisca; Rodrigo-Pendás, Jose Ángel; Noguera-Julian, Antoni; Martínez-Lacasa, Xavier; Tuñez, Maria Victoria; Fernández, Virginia Leiro; Millet, Joan P; Moreno, Antonio; Cobos, Nazaret; Miró, José M; Roldan, Llanos; Orcau, Angels; Andersen, Peter; Caylá, Joan A

    2017-04-01

    Targeted screening and treatment of Mycobacterium tuberculosis infection substantially reduces the risk of developing active tuberculosis. C-Tb (Statens Serum Institute, Copenhagen, Denmark) is a novel specific skin test based on ESAT-6 and CFP10 antigens. We investigated the safety and diagnostic potential of C-Tb compared with established tests in the contact-tracing setting. Negative controls, close contacts, occasional contacts, and patients with active pulmonary tuberculosis were enrolled at 13 centres in Spain. We compared C-Tb with the QuantiFERON-TB Gold In-Tube ([QFT] Qiagen, Hilden, Germany) interferon γ release assay (IGRA) and the purified protein derivative (PPD) RT 23 tuberculin skin test ([TST] Statens Serum Institute). All participants older than 5 years were tested with QFT. Some participants in the negative control group received C-Tb without the TST to test for potential interactions between C-Tb and PPD RT 23. The rest were randomly assigned in blocks of ten and tested with both C-Tb and TST, with five in each block receiving injection of C-Tb in the right arm and the TST in the left arm and five vice versa. The primary and safety analyses were done in all participants randomly assigned to a group who received any test. This trial is registered with ClinicalTrials.gov, number NCT01631266, and with EudraCT, number 2011-005617-36. From July 24, 2012, to Oct 2, 2014, 979 participants were enrolled, of whom 263 were negative controls, 299 were occasional contacts, 316 were close contacts, and 101 were patients with tuberculosis. 970 (99%) participants completed the trial. Induration sizes were similar for C-Tb and TST, but TST positivity was affected by BCG vaccination status. We found a strong positive trend towards C-Tb test positivity with increasing risk of infection, from 3% in negative controls to 16% in occasional contacts, to 43% in close contacts. C-Tb and QFT results were concordant in 785 (94%) of 834 participants aged 5 years and older

  18. Influence of exercise on NA- and Hsp72-induced release of IFNγ by the peritoneal suspension of macrophages and lymphocytes from genetically obese Zucker rats.

    Science.gov (United States)

    Martín-Cordero, L; García, J J; Hinchado, M D; Bote, E; Ortega, E

    2013-03-01

    Regular physical exercise is recognized as a nonpharmacological therapeutic strategy in the treatment of metabolic syndrome, and has been proposed for improving obesity, diabetic status, insulin resistance, and immune response. The aim of the present study was to evaluate the effect of a regular exercise program (treadmill running, 5 days/week for 14 weeks at 35 cm/s for 35 min in the last month) on the release of the pro-inflammatory cytokine interferon gamma (IFNγ) by peritoneal cells (macrophages and lymphocytes) from obese Zucker rats (fa/fa) in response to noradrenaline (NA) and heat shock proteins of 72 kDa (Hsp72), and the possible adaptation due to training for a bout acute exercise (a single session of 25-35 min at 35 cm/s). In healthy (lean Fa/fa) and obese animals, peritoneal cells released greater concentrations of IFNγ in response to Hsp72 and lower concentrations in response to NA. The regular exercise training protocol, evaluated in the obese animals, produced a clear change in the regulation of the release of IFNγ. Peritoneal immune cells from trained animals released more IFNγ in response to NA, but there was a reduction in the release of IFNγ in response to Hsp72. In the obese animals, regular exercise caused a change in the inhibitory effect of NA (which now becomes stimulatory) and the stimulatory effect of Hsp72e (which now becomes inhibitory) in relation to the release of IFNγ. This reflects that Hsp72, induced by the prior release of NA following exercise-induced stress, plays a role in the homeostatic balance of release of IFNγ by peritoneal immune cells in obese animals during exercise.

  19. An ultrafiltration assay for lysyl oxidase

    International Nuclear Information System (INIS)

    Shackleton, D.R.; Hulmes, D.J.

    1990-01-01

    A modification of the original microdistillation assay for lysyl oxidase is described in which Amicon C-10 microconcentrators are used to separate, by ultrafiltration, the 3H-labeled products released from a [4,5-3H]-lysine-labeled elastin substrate. Enzyme activity is determined by scintillation counting of the ultrafiltrate, after subtraction of radioactivity released in the presence of beta-aminopropionitrile, a specific inhibitor of the enzyme. Conditions are described which optimize both the sensitivity and the efficient use of substrate. The assay shows linear inhibition of activity in up to 1 M urea; hence, as the enzyme is normally diluted in the assay, samples in 6 M urea can be assayed directly, without prior dialysis, and corrected for partial inhibition. Comparable results are obtained when enzyme activity is assayed by ultrafiltration or microdistillation. The assay is simple and convenient and, by using disposable containers throughout, it eliminates the need for time-consuming decontamination of radioactive glassware

  20. Mechanism and kinetics of the loss of poorly soluble drugs from liposomal carriers studied by a novel flow field-flow fractionation-based drug release-/transfer-assay

    DEFF Research Database (Denmark)

    Hinna, Askell Hvid; Hupfeld, Stefan; Kuntsche, Judith

    2016-01-01

    Liposomes represent a versatile drug formulation approach e.g. for improving the water-solubility of poorly soluble drugs but also to achieve drug targeting and controlled release. For the latter applications it is essential that the drug remains associated with the liposomal carrier during transit...... in the vascular bed. A range of in vitro test methods has been suggested over the years for prediction of the release of drug from liposomal carriers. The majority of these fail to give a realistic prediction for poorly water-soluble drugs due to the intrinsic tendency of such compounds to remain associated...... the amount of drug remaining associated with the liposomal drug carrier as well as that transferred to the acceptor liposomes at distinct times of incubation, boththe kinetics of drug transfer and release to the water phase could be established for the model drug p-THPP (5,10,15,20-tetrakis(4-hydroxyphenyl...

  1. Insecticidal and sterilizing effect of Olyset Duo (R), a permethrin and pyriproxyfen mixture net against pyrethroid-susceptible and -resistant strains of Anopheles gambiae s.s. : a release-recapture assay in experimental huts

    OpenAIRE

    Djènontin, A.; Alou, L. P. A.; Koffi, A.; Zogo, B.; Duarte, E.; N'Guessan, R.; Moiroux, Nicolas; Pennetier, Cédric

    2015-01-01

    In the context of the widespread distribution of pyrethroid resistance among malaria vectors, we did a release-recapture trial in experimental huts to investigate the insecticidal and sterilizing effects of a novel long-lasting net (LN), Olyset (R) Duo, incorporating a mixture of permethrin (PER) and the insect growth regulator (IGR), pyriproxyfen (PPF). An LN containing PPF alone and a classic Olyset (R) Net were tested in parallel as positive controls. The effect of progressive number of ho...

  2. Role of interferon-gamma in the pathogenesis of LCMV-induced meningitis: unimpaired leucocyte recruitment, but deficient macrophage activation in interferon-gamma knock-out mice

    DEFF Research Database (Denmark)

    Nansen, A; Christensen, Jan Pravsgaard; Röpke, C

    1998-01-01

    , a viral peptide could also elicit a T cell mediated inflammatory response in virus-primed IFN-gamma knock-out mice, indicating that redundancy of this cytokine as a proinflammatory mediator is not restricted to inflammatory reactions triggered by an active infection. Thus, T cell mediated inflammation may...

  3. Insecticidal and sterilizing effect of Olyset Duo®, a permethrin and pyriproxyfen mixture net against pyrethroid-susceptible and -resistant strains of Anopheles gambiae s.s.: a release-recapture assay in experimental huts

    Directory of Open Access Journals (Sweden)

    Djènontin Armel

    2015-01-01

    Full Text Available In the context of the widespread distribution of pyrethroid resistance among malaria vectors, we did a release-recapture trial in experimental huts to investigate the insecticidal and sterilizing effects of a novel long-lasting net (LN, Olyset® Duo, incorporating a mixture of permethrin (PER and the insect growth regulator (IGR, pyri-proxyfen (PPF. An LN containing PPF alone and a classic Olyset® Net were tested in parallel as positive controls. The effect of progressive number of holes (6, 30, or 150 that may accrue in nets over time was simulated. We used two laboratory Anopheles gambiae s.s. strains: the susceptible Kisumu strain and the pyrethroid-resistant VK-Per strain having solely kdr as resistance mechanism. The effect of these nets on the reproductive success of blood-fed females that survived the different LNs conditions was recorded. Regardless of the mosquito strain, the LNs containing PPF alone with as many as 30 holes drastically reduced the number of eggs laid by females succeeding in feeding, i.e. fecundity by 98% and egg hatching rate (fertility by 93% relative to untreated control net. Very few of the resistant females blood fed and survived under the Olyset® Duo with similar number of holes (up to 30 but of these few, the inhibition of reproductive success was 100%. There was no evidence that the Olyset® Duo LN with 150 holes impacted fecundity or fertility of the resistant colony. The efficacy of Olyset® Duo is encouraging and clearly illustrates that this new net might be a promising tool for malaria transmission control and resistance management.

  4. Insecticidal and sterilizing effect of Olyset Duo®, a permethrin and pyriproxyfen mixture net against pyrethroid-susceptible and -resistant strains of Anopheles gambiae s.s.: a release-recapture assay in experimental huts.

    Science.gov (United States)

    Djènontin, Armel; Ahoua Alou, Ludovic P; Koffi, Alphonsine; Zogo, Barnabas; Duarte, Elves; N'Guessan, Raphael; Moiroux, Nicolas; Pennetier, Cédric

    2015-01-01

    In the context of the widespread distribution of pyrethroid resistance among malaria vectors, we did a release-recapture trial in experimental huts to investigate the insecticidal and sterilizing effects of a novel long-lasting net (LN), Olyset® Duo, incorporating a mixture of permethrin (PER) and the insect growth regulator (IGR), pyri-proxyfen (PPF). An LN containing PPF alone and a classic Olyset® Net were tested in parallel as positive controls. The effect of progressive number of holes (6, 30, or 150) that may accrue in nets over time was simulated. We used two laboratory Anopheles gambiae s.s. strains: the susceptible Kisumu strain and the pyrethroid-resistant VK-Per strain having solely kdr as resistance mechanism. The effect of these nets on the reproductive success of blood-fed females that survived the different LNs conditions was recorded. Regardless of the mosquito strain, the LNs containing PPF alone with as many as 30 holes drastically reduced the number of eggs laid by females succeeding in feeding, i.e. fecundity by 98% and egg hatching rate (fertility) by 93% relative to untreated control net. Very few of the resistant females blood fed and survived under the Olyset® Duo with similar number of holes (up to 30) but of these few, the inhibition of reproductive success was 100%. There was no evidence that the Olyset® Duo LN with 150 holes impacted fecundity or fertility of the resistant colony. The efficacy of Olyset® Duo is encouraging and clearly illustrates that this new net might be a promising tool for malaria transmission control and resistance management. © A. Djènontin et al., published by EDP Sciences, 2015.

  5. Interaction with extracellular matrix proteins influences Lsh/Ity/Bcg (candidate Nramp) gene regulation of macrophage priming/activation for tumour necrosis factor-alpha and nitrite release.

    Science.gov (United States)

    Formica, S; Roach, T I; Blackwell, J M

    1994-05-01

    The murine resistance gene Lsh/Ity/Bcg regulates activation of macrophages for tumour necrosis factor-alpha (TNF-alpha)-dependent production of nitric oxide mediating antimicrobial activity against Leishmania, Salmonella and Mycobacterium. As Lsh is differentially expressed in macrophages from different tissue sites, experiments were performed to determine whether interaction with extracellular matrix (ECM) proteins would influence the macrophage TNF-alpha response. Plating of bone marrow-derived macrophages onto purified fibrinogen or fibronectin-rich L929 cell-derived matrices, but not onto mannan, was itself sufficient to stimulate TNF-alpha release, with significantly higher levels released from congenic B10.L-Lshr compared to C57BL/10ScSn (Lshs) macrophages. Only macrophages plated onto fibrinogen also released measurable levels of nitrites, again higher in Lshr compared to Lshs macrophages. Addition of interferon-gamma (IFN-gamma), but not bacterial lipopolysaccharide or mycobacterial lipoarabinomannan, as a second signal enhanced the TNF-alpha and nitrite responses of macrophages plated onto fibrinogen, particularly in the Lshr macrophages. Interaction with fibrinogen and fibronectin also primed macrophages for an enhanced TNF-alpha response to leishmanial parasites, but this was only translated into enhanced nitrite responses in the presence of IFN-gamma. In these experiments, Lshr macrophages remained superior in their TNF-alpha responses throughout, but to a degree which reflected the magnitude of the difference observed on ECM alone. Hence, the specificity for the enhanced TNF-alpha responses of Lshr macrophages lay in their interaction with fibrinogen and fibronectin ECM, while a differential nitrite response was only observed with fibrinogen and/or IFN-gamma. The results are discussed in relation to the possible function of the recently cloned candidate gene Nramp, which has structural identity to eukaryote transporters and an N-terminal cytoplasmic

  6. Methane release

    International Nuclear Information System (INIS)

    Seifert, M.

    1999-01-01

    The Swiss Gas Industry has carried out a systematic, technical estimate of methane release from the complete supply chain from production to consumption for the years 1992/1993. The result of this survey provided a conservative value, amounting to 0.9% of the Swiss domestic output. A continuation of the study taking into account new findings with regard to emission factors and the effect of the climate is now available, which provides a value of 0.8% for the target year of 1996. These results show that the renovation of the network has brought about lower losses in the local gas supplies, particularly for the grey cast iron pipelines. (author)

  7. Microbead agglutination based assays

    KAUST Repository

    Kodzius, Rimantas

    2013-01-21

    We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microbeads in the presence of a specific analyte thus enabling the macroscopic observation. Such tests are most often used to explore antibody-antigen reactions. Agglutination has been used for protein assays using a biotin/streptavidin system as well as a hybridization based assay. The agglutination systems are prone to selftermination of the linking analyte, prone to active site saturation and loss of agglomeration at high analyte concentrations. We investigated the molecular target/ligand interaction, explaining the common agglutination problems related to analyte self-termination, linkage of the analyte to the same bead instead of different microbeads. We classified the agglutination process into three kinds of assays: a two- component assay, a three-component assay and a stepped three- component assay. Although we compared these three kinds of assays for recognizing DNA and protein molecules, the assay can be used for virtually any molecule, including ions and metabolites. In total, the optimized assay permits detecting analytes with high sensitivity in a short time, 5 min, at room temperature. Such a system is appropriate for POC testing.

  8. Active case finding of tuberculosis in Europe: a Tuberculosis Network European Trials Group (TBNET) survey

    DEFF Research Database (Denmark)

    Bothamley, G.H.; Ditiu, L.; Migliori, G.B.

    2008-01-01

    countries do not, even if the chest radiograph suggests tuberculosis. Nearly all countries (49 out of 50) use tuberculin skin testing (TST); 27 (54%) out of 50 countries also perform chest radiography irrespective of the TST result. Interpretation of the TST varies widely. All countries use 6-9 months...... of isoniazid for treatment of LTBI, with an estimated median (range) uptake of 55% (5-92.5%). Symptoms and sputum examination could be used more widely when screening for active tuberculosis. Treatment of latent tuberculosis infection might be better focused by targeted use of interferon-gamma release assays...

  9. Radioreceptor opioid assay

    International Nuclear Information System (INIS)

    Miller, R.J.; Chang, K.-J.

    1981-01-01

    A radioreceptor assay is described for assaying opioid drugs in biological fluids. The method enables the assay of total opioid activity, being specific for opioids as a class but lacking specificity within the class. A radio-iodinated opioid and the liquid test sample are incubated with an opiate receptor material. The percentage inhibition of the binding of the radio-iodinated compound to the opiate receptor is calculated and the opioid activity of the test liquid determined from a standard curve. Examples of preparing radio-iodinated opioids and assaying opioid activity are given. A test kit for the assay is described. Compared to other methods, this assay is cheap, easy and rapid. (U.K.)

  10. Absolute nuclear material assay

    Science.gov (United States)

    Prasad, Manoj K [Pleasanton, CA; Snyderman, Neal J [Berkeley, CA; Rowland, Mark S [Alamo, CA

    2010-07-13

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  11. The glucose oxidase-peroxidase assay for glucose

    Science.gov (United States)

    The glucose oxidase-peroxidase assay for glucose has served as a very specific, sensitive, and repeatable assay for detection of glucose in biological samples. It has been used successfully for analysis of glucose in samples from blood and urine, to analysis of glucose released from starch or glycog...

  12. Evaluation of the Erythrocyte Malondialdehyde (MDA) Release Assay.

    Science.gov (United States)

    1987-01-01

    MDA vs. Cholesterol A It.M U Control indicated that there was alsc: a p-sit ive .. r i Piat i r, 0.58) between cholestetol an’] I I asma ::im E...N Engl J Med 310, 1209-1212(1984). 58. Hess AF. Involvement of the blood and blood vessels in infantile scurvy. Proc Soc Exp Biol Med 11, 130-132

  13. Interferon gamma/NADPH oxidase defense system in immunity and cancer

    Czech Academy of Sciences Publication Activity Database

    Hodný, Zdeněk; Reiniš, Milan; Hubáčková, Soňa; Vašicová, Pavla; Bartek, Jiří

    2016-01-01

    Roč. 5, č. 2 (2016), č. článku e1080416. ISSN 2162-402X Institutional support: RVO:68378050 ; RVO:61388971 Keywords : Cellular senescence * DNA damage response * oxidative stress * tumor growth factor beta * tumor immunosurveillance Subject RIV: EB - Genetics ; Molecular Biology; EC - Immunology (MBU-M) Impact factor: 7.719, year: 2016

  14. cd4+ and cd8+ t-cells expressing interferon gamma in active ...

    African Journals Online (AJOL)

    USER

    2017-10-25

    Oct 25, 2017 ... tuberculosis live in South East Asia and West Pacific with three hundred and ... is ranked fourth (410-520 thousand) after India (2-2.3 million), China (0.9-1.1 million), and Nigeria (340-880 ... Sputum culture status in the second.

  15. Systemic and local interferon-gamma production following Mycobacterium ulcerans infection

    NARCIS (Netherlands)

    Schipper, H. S.; Rutgers, B.; Huitema, M. G.; Etuaful, S. N.; Westenbrink, B. D.; Limburg, P. C.; Timens, W.; van der Werf, T. S.

    2007-01-01

    Buruli ulcer disease (BUD) is an emerging predominantly tropical disease caused by Mycobacterium ulcerans. The initial pre-ulcerative skin lesion often breaks down into an ulcer with undermined edges. Healing is common but may require considerable time, and scarring often results in functional

  16. Interleukin-4 (IL-4) and Interferon-Gamma (IFN-gamma) in pregnant ...

    African Journals Online (AJOL)

    Background and Objective:- To assess if gestational factors affect the resistance of C57BL/6 mice to L. major infection, this study determined the levels of IL-4 and IFN-gamma in popliteal lymph node cells of pregnant C57BL/6 mice infected with L. major at 16 hours, 5 days-, 10 days- and 15 days- post plug by PCR, ELISA ...

  17. The Role of the Interferon-Gamma-Jak/STAT Pathway in Rheumatoid Arthritis

    Science.gov (United States)

    2017-09-01

    including suggestions for reducing this burden to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and...cytometry the T cell populations, the B cell populations and monocytes. Beginning Month 11, we will begin interrogating these cell populations for the... interrogate IFN-γ induced SAT1 activation along with the activation of STAT3 and STAT5 over a range of stimulation time- points. This approach is feasible

  18. The effects of interferon-gamma (INF-γ) on the expression of insulin ...

    African Journals Online (AJOL)

    ) on insulin-like growth factor 1(IGF-1) and its effect on pregnancy, the dynamic expression of mRNA and cellular localization of IGF-1 protein in the early pregnant (Day 9) rats after the injection of different doses of IFN-γ, their ovaries and ...

  19. Human keratinocytes produce the complement inhibitor factor H: synthesis is regulated by interferon-gamma

    NARCIS (Netherlands)

    Timár, Krisztina K.; Pasch, Marcel C.; van den Bosch, Norbert H. A.; Jarva, Hanna; Junnikkala, Sami; Meri, Seppo; Bos, Jan D.; Asghar, Syed S.

    2006-01-01

    Locally synthesized complement is believed to play an important role in host defense and inflammation at organ level. In the epidermis, keratinocytes have so far been shown to synthesize two complement components, C3 and factor B. Here, we studied the synthesis of factor H by human keratinocytes. We

  20. Polymorphisms in an interferon-gamma receptor-1 gene marker and susceptibility to periodontitis

    NARCIS (Netherlands)

    Fraser, DA; Loos, BG; Boman, U; van Winkelhoff, AJ; van der Velden, U; Schenck, K; Dembic, Z

    2003-01-01

    Chronic marginal periodontitis is an inflammatory condition in which the supporting tissues of the teeth are destroyed. Interferon (IFN)-gamma is a cytokine that plays a pivotal role in the defense against infection, and mutations in the gene coding for the ligand binding chain (alpha, RI) of the

  1. Diminished interferon-gamma production and responsiveness after endotoxin administration to healthy humans

    NARCIS (Netherlands)

    Weijer, Sebastiaan; Lauw, Fanny N.; Branger, Judith; van den Blink, Bernt; van der Poll, Tom

    2002-01-01

    To obtain insight in the capacity of the lipopolysaccharide (LPS)-tolerant host to produce interferon (IFN)-gamma and to respond to this cytokine, whole blood was obtained from healthy humans before and 4 h after intravenous injection of LPS (4 ng/kg) and stimulated ex vivo. LPS exposure in vivo

  2. Intracellular fate of recombinant human interferon-gamma (rIFN) in U937 cells

    International Nuclear Information System (INIS)

    Finbloom, D.S.

    1986-01-01

    After IFN binds to specific receptors on macrophages, both modulation of surface molecules and induction of microbicidal and tumoricidal activity occurs 24-48 hr later. Since the intracellular events required to insure these responses are poorly defined, the fate of radiolabeled rIFN in U937 cells was examined. Endocytosis was determined by exposing cells to pH 2.5 to allow rIFN to dissociate leaving only intracellular ligand. Degradation was measured as trichloroacetic acid soluble radioactivity in the media. Of the 4-5000 molecules of rIFN that specifically and saturably (at 300 U/ml) bound at 4 0 C, 40% dissociated during 15-30 min after cells were warmed to 37 0 C. However, if cells were continuously exposed at 37 0 C to lower levels of rIFN (60-100 U/ml), 30-40% of those molecules capable of binding to the cell at that concentration were internalized. Furthermore, 60% of the molecules were degraded during 3-4 hr of additional culture. Since exposure of cells to chloroquine and monensin resulted in only partial inhibition of degradation (75% and 43%, respectively), there may also be degradation within endosomes or on the cell following binding to its receptor. Soon thereafter, degradation products are measurable. Since many biological responses require prolonged incubation with the molecule, intracellular processing of IFN may be important for expression of these effects

  3. Interferon-gamma impairs proliferation of hematopoietic stem cells in mice

    NARCIS (Netherlands)

    de Bruin, Alexander M.; Demirel, Özlem; Hooibrink, Berend; Brandts, Christian H.; Nolte, Martijn A.

    2013-01-01

    Balancing the processes of hematopoietic stem cell (HSC) differentiation and self-renewal is critical for maintaining a lifelong supply of blood cells. The bone marrow (BM) produces a stable output of newly generated cells, but immunologic stress conditions inducing leukopenia increase the demand

  4. Antigen-Specific Interferon-Gamma Responses and Innate Cytokine Balance in TB-IRIS

    NARCIS (Netherlands)

    Goovaerts, Odin; Jennes, Wim; Massinga-Loembé, Marguerite; Ceulemans, Ann; Worodria, William; Mayanja-Kizza, Harriet; Colebunders, Robert; Kestens, Luc; Loembé, Marguerite Massinga; Mayanja, Harriet; Mascart, Francoise; van den Bergh, Rafael; Locht, Camille; Reiss, Peter; Cobelens, Frank; Ondoa, Pascale; Pakker, Nadine; Mugerwa, Roy

    2014-01-01

    Background: Tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) remains a poorly understood complication in HIV-TB patients receiving antiretroviral therapy (ART). TB-IRIS could be associated with an exaggerated immune response to TB-antigens. We compared the recovery of

  5. Antigen-Specific Interferon-Gamma Responses and Innate Cytokine Balance in TB-IRIS

    Science.gov (United States)

    Goovaerts, Odin; Jennes, Wim; Massinga-Loembé, Marguerite; Ceulemans, Ann; Worodria, William; Mayanja-Kizza, Harriet; Colebunders, Robert; Kestens, Luc

    2014-01-01

    Background Tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) remains a poorly understood complication in HIV-TB patients receiving antiretroviral therapy (ART). TB-IRIS could be associated with an exaggerated immune response to TB-antigens. We compared the recovery of IFNγ responses to recall and TB-antigens and explored in vitro innate cytokine production in TB-IRIS patients. Methods In a prospective cohort study of HIV-TB co-infected patients treated for TB before ART initiation, we compared 18 patients who developed TB-IRIS with 18 non-IRIS controls matched for age, sex and CD4 count. We analyzed IFNγ ELISpot responses to CMV, influenza, TB and LPS before ART and during TB-IRIS. CMV and LPS stimulated ELISpot supernatants were subsequently evaluated for production of IL-12p70, IL-6, TNFα and IL-10 by Luminex. Results Before ART, all responses were similar between TB-IRIS patients and non-IRIS controls. During TB-IRIS, IFNγ responses to TB and influenza antigens were comparable between TB-IRIS patients and non-IRIS controls, but responses to CMV and LPS remained significantly lower in TB-IRIS patients. Production of innate cytokines was similar between TB-IRIS patients and non-IRIS controls. However, upon LPS stimulation, IL-6/IL-10 and TNFα/IL-10 ratios were increased in TB-IRIS patients compared to non-IRIS controls. Conclusion TB-IRIS patients did not display excessive IFNγ responses to TB-antigens. In contrast, the reconstitution of CMV and LPS responses was delayed in the TB-IRIS group. For LPS, this was linked with a pro-inflammatory shift in the innate cytokine balance. These data are in support of a prominent role of the innate immune system in TB-IRIS. PMID:25415590

  6. Latent tuberculosis in HIV positive, diagnosed by the M. tuberculosis specific interferon-gamma test

    DEFF Research Database (Denmark)

    Brock, Inger; Ruhwald, Morten; Lundgren, Bettina

    2006-01-01

    Although tuberculosis (TB) is a minor problem in Denmark, severe and complicated cases occur in HIV positive. Since the new M. tuberculosis specific test for latent TB, the QuantiFERON-TB In-Tube test (QFT-IT) became available the patients in our clinic have been screened for the presence of latent...

  7. Latent tuberculosis in HIV positive, diagnosed by the M. tuberculosis specific interferon-gamma test

    DEFF Research Database (Denmark)

    Brock, Inger; Ruhwald, Morten; Lundgren, Bettina

    2006-01-01

    BACKGROUND: Although tuberculosis (TB) is a minor problem in Denmark, severe and complicated cases occur in HIV positive. Since the new M. tuberculosis specific test for latent TB, the QuantiFERON-TB In-Tube test (QFT-IT) became available the patients in our clinic have been screened...

  8. Interferon gamma/NADPH oxidase defence system in immunity and cancer

    Czech Academy of Sciences Publication Activity Database

    Hodný, Zdeněk; Reiniš, Milan; Hubáčková, Soňa; Vašicová, Pavla; Bartek, Jiří

    -, 01 Sep (2015) ISSN 2162-4011 Institutional support: RVO:68378050 ; RVO:61388971 Keywords : IFNγ * NADPH oxidase * immunity * cancer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.266, year: 2014

  9. Distinct interferon-gamma and interleukin-9 expression in cutaneous and oral lichen planus.

    Science.gov (United States)

    Weber, B; Schlapbach, C; Stuck, M; Simon, H-U; Borradori, L; Beltraminelli, H; Simon, D

    2017-05-01

    Cutaneous (CLP) and oral lichen planus (OLP) as the main subtypes of lichen planus (LP) present with different clinical manifestation and disease course, although their histopathologic features such as the band-like lymphocyte infiltrate and keratinocyte apoptosis are similar. So far, the underlying cellular and molecular mechanisms remain poorly understood. The aim of this study was to characterize and compare the in situ cellular infiltrates, cytokine expression profiles and apoptosis markers in CLP and OLP. Using immunofluorescence staining and laser scanning microscopy, we evaluated the cellular infiltrate (CD1a, CD3, CD4, CD8, CD21, CD57, CD123), cytokine expression (interleukin (IL)-1, IL-6, IL-9, IL-10, IL-17, IL-22, IL-23, tumour necrosis factor-α, transforming growth factor-β, interferon (IFN)-γ), and apoptosis markers (Fas, Fas ligand, cleaved caspase-3, TUNEL) of 21 anonymized biopsy specimens of LP (11 CLP, 10 OLP). Among infiltrating cells mainly T cells and natural killer (NK) cells as well as plasmacytoid dendritic cells (DC) were observed. A predominance of CD8+ T cells was noted in OLP. In both CLP and OLP, T helper (Th)1, Th9, Th17, and Th22-type cytokines were expressed. The expression of IL-9, IFN-γ and IL-22 was higher in CLP compared to that of OLP (P = 0.0165; P = 0.0016; P = 0.052 respectively). Expression of Fas and Fas ligand as well as cleaved caspase-3-positive cells was observed in the epithelium of all LP samples. The cell and cytokine patterns of CLP and OLP were partially distinct and generally resembled those reported for autoimmune diseases. The presence of CD8+ and NK cells as well as Fas/Fas ligand expression suggested that various pathways involved in keratinocyte apoptosis are relevant for LP. These results might help to establish targeted therapies for LP. © 2016 European Academy of Dermatology and Venereology.

  10. Endogenous Locus Reporter Assays.

    Science.gov (United States)

    Liu, Yaping; Hermes, Jeffrey; Li, Jing; Tudor, Matthew

    2018-01-01

    Reporter gene assays are widely used in high-throughput screening (HTS) to identify compounds that modulate gene expression. Traditionally a reporter gene assay is built by cloning an endogenous promoter sequence or synthetic response elements in the regulatory region of a reporter gene to monitor transcriptional activity of a specific biological process (exogenous reporter assay). In contrast, an endogenous locus reporter has a reporter gene inserted in the endogenous gene locus that allows the reporter gene to be expressed under the control of the same regulatory elements as the endogenous gene, thus more accurately reflecting the changes seen in the regulation of the actual gene. In this chapter, we introduce some of the considerations behind building a reporter gene assay for high-throughput compound screening and describe the methods we have utilized to establish 1536-well format endogenous locus reporter and exogenous reporter assays for the screening of compounds that modulate Myc pathway activity.

  11. A sensitive assay for Staphylococcus aureus nucleases

    Energy Technology Data Exchange (ETDEWEB)

    Kohli, J K; Vakil, B V; Patil, M S; Pandey, V N; Pradhan, D S [Bhabha Atomic Reserach Centre, Bombay (India). Biochemistry Div.

    1989-10-01

    A sensitive assay for staphylococcal nuclease involving incubation of the enzyme sample with heat-denatured ({sup 3}H) thymidine labelled DNA from E.coli, precipitation with trichloroacetic acid and measurement of the radioactivity of acid-soluble nucleotides released has been developed. The assay is sensitive enough to be used for comparing the levels of nucleases elaborated by different strains of S. aureus as well as for determining the extent of contamination of S. aureus in food and water samples even at levels at which the conventional spectrophotometric and toluidine blue-DNA methods are totally inadequate. (author). 26 refs., 3 figs ., 3 tabs.

  12. Solid phase assays

    International Nuclear Information System (INIS)

    Reese, M.G.; Johnson, L.R.; Ransom, D.K.

    1980-01-01

    In a solid phase assay for quantitative determination of biological and other analytes, a sample such as serum is contacted with a receptor for the analyte being assayed, the receptor being supported on a solid support. No tracer for the analyte is added to the sample before contacting with the receptor; instead the tracer is contacted with the receptor after unbound analyte has been removed from the receptor. The assay can be otherwise performed in a conventional manner but can give greater sensitivity. (author)

  13. Relationship between the radioisotopic footpad assay and other immunological assays in tumor bearing rats

    International Nuclear Information System (INIS)

    Mizushima, Yutaka; Takeichi, Noritoshi; Minami, Akio; Kasai, Masaharu; Itaya, Toshiyuki

    1981-01-01

    KMT-17, a fibrosarcoma induced by 3-methylcholanthrene in a WKA rat, is a sensitive tumor to various kinds of immunological assays and is a suitable model tumor for the study of the immune status in tumor bearing hosts. The antitumor immune response of KMT-17 bearing rats was studied by a radioisotopic footpad assay (FPA) in comparison with other in vivo and in vitro assays. Delayed hypersensitivity to tumor antigens measured by the FPA was observed from the 8th day after transplantation of KMT-17 cells, reached a peak on the 12 - 15th day, and then declined in the late stage on the 17th day. The kinetics of the FPA correlated well with those of an in vivo Winn assay and of an in vitro lymphocyte cytotoxicity assay ( 51 Cr-release assay). The appearance of an antitumor antibody detected by a complement dependent cytotoxicity test also correlated well with the kinetics of the FPA. A growth inhibition assay (GIA) for non-specific cell-mediated immunity also showed similar kinetics to that of the FPA. The delayed hypersensitivity footpad reaction to tumor cell extracts measured by this FPA was tumor-specific. These results suggest that the FPA is a simple and reliable in vivo assay for evaluating antitumor immunity in tumor bearing hosts. (author)

  14. Radioimmune assay of human platelet prostaglandin synthetase

    International Nuclear Information System (INIS)

    Roth, G.J.; Machuga, E.T.

    1982-01-01

    Normal platelet function depends, in part, on platelet PG synthesis. PG synthetase (cyclo-oxygenase) catalyzes the first step in PG synthesis, the formation of PGH 2 from arachidonic acid. Inhibition of the enzyme by ASA results in an abnormality in the platelet release reaction. Patients with pparent congenital abnormalities in the enzyme have been described, and the effects have been referred to as ''aspirin-like'' defects of the platelet function. These patients lack platelet PG synthetase activity, but the actual content of PG synthetase protein in these individuals' platelets is unknown. Therefore an RIA for human platelet PG synthetase would provide new information, useful in assessing the aspirin-like defects of platelet function. An RIA for human platelet PG synthetase is described. The assay utilizes a rabbit antibody directed against the enzyme and [ 125 I]-labelled sheep PG synthetase as antigen. The human platelet enzyme is assayed by its ability to inhibit precipitation of the [ 125 I]antigen. The assay is sensitive to 1 ng of enzyme. By the immune assay, human platelets contain approximately 1200 ng of PG synethetase protein per 1.5 mg of platelet protein (approximately 10 9 platelets). This content corresponds to 10,000 enzyme molecules per platelet. The assay provides a rapid and convenient assay for the human platelet enzyme, and it can be applied to the assessment of patients with apparent platelet PG synthetase (cyclo-oxygenase) deficiency

  15. Factor IX assay

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003679.htm Factor IX assay To use the sharing features on ... M. is also a founding member of Hi-Ethics and subscribes to the principles of the Health ...

  16. Factor VIII assay

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003678.htm Factor VIII assay To use the sharing features on ... M. is also a founding member of Hi-Ethics and subscribes to the principles of the Health ...

  17. Factor II assay

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003674.htm Factor II assay To use the sharing features on ... M. is also a founding member of Hi-Ethics and subscribes to the principles of the Health ...

  18. Factor VII assay

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003676.htm Factor VII assay To use the sharing features on ... M. is also a founding member of Hi-Ethics and subscribes to the principles of the Health ...

  19. Microbead agglutination based assays

    KAUST Repository

    Kodzius, Rimantas; Castro, David; Foulds, Ian G.; Parameswaran, Ash M.; Sumanpreet, K. Chhina

    2013-01-01

    We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microbeads in the presence of a specific analyte thus enabling

  20. Assay method and compositions

    International Nuclear Information System (INIS)

    1977-01-01

    Methods are described for measuring catecholamine levels in human and animal body fluids and tissues using the catechol-O-methyl-transferase (COMT) radioassay. The assay involves incubating the biological sample with COMT and the tritiated methyl donor, S-adenosyl-L-methionine( 3 H)-methyl. The O-methylated ( 3 H) epinephrine and/or norepinephrine are extracted and oxidised to vanillin- 3 H which in turn is extracted and its radioactivity counted. When analysing dopamine levels the assay is extended by vanillin- 3 H and raising the pH of the aqueous periodate phase from which O-methylated ( 3 H) dopamine is extracted and counted. The assay may be modified depending on whether measurements of undifferentiated total endogenous catecholamine levels or differential analyses of the catecholamine levels are being performed. The sensitivity of the assay can be as low as 5 picograms for norepinephrine and epinephrine and 12 picograms for dopamine. The assemblance of the essential components of the assay into a kit for use in laboratories is also described. (U.K.)

  1. Rover waste assay system

    Energy Technology Data Exchange (ETDEWEB)

    Akers, D.W.; Stoots, C.M.; Kraft, N.C.; Marts, D.J. [Idaho National Engineering Lab., Idaho Falls, ID (United States)

    1997-11-01

    The Rover Waste Assay System (RWAS) is a nondestructive assay system designed for the rapid assay of highly-enriched {sup 235}U contaminated piping, tank sections, and debris from the Rover nuclear rocket fuel processing facility at the Idaho Chemical Processing Plant. A scanning system translates a NaI(Tl) detector/collimator system over the structural components where both relative and calibrated measurements for {sup 137}Cs are made. Uranium-235 concentrations are in operation and is sufficiently automated that most functions are performed by the computer system. These functions include system calibration, problem identification, collimator control, data analysis, and reporting. Calibration of the system was done through a combination of measurements on calibration standards and benchmarked modeling. A description of the system is presented along with the methods and uncertainties associated with the calibration and analysis of the system for components from the Rover facility. 4 refs., 2 figs., 4 tabs.

  2. Rover waste assay system

    International Nuclear Information System (INIS)

    Akers, D.W.; Stoots, C.M.; Kraft, N.C.; Marts, D.J.

    1997-01-01

    The Rover Waste Assay System (RWAS) is a nondestructive assay system designed for the rapid assay of highly-enriched 235 U contaminated piping, tank sections, and debris from the Rover nuclear rocket fuel processing facility at the Idaho Chemical Processing Plant. A scanning system translates a NaI(Tl) detector/collimator system over the structural components where both relative and calibrated measurements for 137 Cs are made. Uranium-235 concentrations are in operation and is sufficiently automated that most functions are performed by the computer system. These functions include system calibration, problem identification, collimator control, data analysis, and reporting. Calibration of the system was done through a combination of measurements on calibration standards and benchmarked modeling. A description of the system is presented along with the methods and uncertainties associated with the calibration and analysis of the system for components from the Rover facility. 4 refs., 2 figs., 4 tabs

  3. Radioreceptor assay for insulin

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Kazuo [Tokyo Univ. (Japan). Faculty of Medicine

    1975-04-01

    Radioreceptor assay of insulin was discussed from the aspects of the measuring method, its merits and problems to be solved, and its clinical application. Rat liver 10 x g pellet was used as receptor site, and enzymatic degradation of insulin by the system contained in this fraction was inhibited by adding 1 mM p-CMB. /sup 125/I-labelled porcine insulin was made by lactoperoxidase method under overnight incubation at 4/sup 0/C and later purification by Sephadex G-25 column and Whatman CF-11 cellulose powder. Dog pancreatic vein serum insulin during and after the glucose load was determined by radioreceptor assay and radioimmunoassay resulting that both measurements accorded considerably. Radioreceptor assay would clarify the pathology of disorders of glucose metabolism including diabetes.

  4. Clonogenic assay: adherent cells.

    Science.gov (United States)

    Rafehi, Haloom; Orlowski, Christian; Georgiadis, George T; Ververis, Katherine; El-Osta, Assam; Karagiannis, Tom C

    2011-03-13

    The clonogenic (or colony forming) assay has been established for more than 50 years; the original paper describing the technique was published in 1956. Apart from documenting the method, the initial landmark study generated the first radiation-dose response curve for X-ray irradiated mammalian (HeLa) cells in culture. Basically, the clonogenic assay enables an assessment of the differences in reproductive viability (capacity of cells to produce progeny; i.e. a single cell to form a colony of 50 or more cells) between control untreated cells and cells that have undergone various treatments such as exposure to ionising radiation, various chemical compounds (e.g. cytotoxic agents) or in other cases genetic manipulation. The assay has become the most widely accepted technique in radiation biology and has been widely used for evaluating the radiation sensitivity of different cell lines. Further, the clonogenic assay is commonly used for monitoring the efficacy of radiation modifying compounds and for determining the effects of cytotoxic agents and other anti-cancer therapeutics on colony forming ability, in different cell lines. A typical clonogenic survival experiment using adherent cells lines involves three distinct components, 1) treatment of the cell monolayer in tissue culture flasks, 2) preparation of single cell suspensions and plating an appropriate number of cells in petri dishes and 3) fixing and staining colonies following a relevant incubation period, which could range from 1-3 weeks, depending on the cell line. Here we demonstrate the general procedure for performing the clonogenic assay with adherent cell lines with the use of an immortalized human keratinocyte cell line (FEP-1811). Also, our aims are to describe common features of clonogenic assays including calculation of the plating efficiency and survival fractions after exposure of cells to radiation, and to exemplify modification of radiation-response with the use of a natural antioxidant

  5. Scintillation proximity assay

    International Nuclear Information System (INIS)

    Hart, H.

    1980-01-01

    In a method of immunological assay two different classes of particles which interact at short distances to produce characteristic detectable signals are employed in a modification of the usual latex fixation test. In one embodiment an aqueous suspension of antigen coated tritiated latex particles (LH) and antigen coated polystyrene scintillant particles (L*) is employed to assay antibody in the aqueous medium. The amount of (LH) (L*) dimer formation and higher order aggregation induced and therefore the concentration of antibody (or antigen) present which caused the aggregation can be determined by using standard liquid scintillation counting equipment. (author)

  6. Assays for calcitonin receptors

    International Nuclear Information System (INIS)

    Teitelbaum, A.P.; Nissenson, R.A.; Arnaud, C.D.

    1985-01-01

    The assays for calcitonin receptors described focus on their use in the study of the well-established target organs for calcitonin, bone and kidney. The radioligand used in virtually all calcitonin binding studies is 125 I-labelled salmon calcitonin. The lack of methionine residues in this peptide permits the use of chloramine-T for the iodination reaction. Binding assays are described for intact bone, skeletal plasma membranes, renal plasma membranes, and primary kidney cell cultures of rats. Studies on calcitonin metabolism in laboratory animals and regulation of calcitonin receptors are reviewed

  7. Evaluation of a MTT assay in measurement of radiosensitizing effect

    International Nuclear Information System (INIS)

    Higuchi, Keiko; Mitsuhashi, Norio; Saitoh, Jun-ichi; Maebayashi, Katsuya; Sakurai, Hideyuki; Akimoto, Tetsuo; Niibe, Hideo

    1999-01-01

    The usefulness of a MTT assay by measuring the radiosensitizing effect of caffeine on rat yolk sac tumor cell line with a mutant-type p53 in vitro was evaluated. A rat yolk sac tumor cell line with a mutant-type p53, NMT-1R, was used in this study. The radiosensitivity of NMT-1R with or without caffeine was measured with a MTT assay. The results were compared with those by a clonogenic assay. Caffeine at a concentration of 2.0 mM which released radiation-induced G 2 block demonstrated a radiosensitizing effect, but caffeine at a concentration of 0.5 mM did not. The radiosensitizing effect of caffeine measured by a MTT assay correlated with that measured by a clonogenic assay. A MTT assay was useful to measure radiosensitivity and/or a radiosensitizing effect in vitro. (author)

  8. News/Press Releases

    Data.gov (United States)

    Office of Personnel Management — A press release, news release, media release, press statement is written communication directed at members of the news media for the purpose of announcing programs...

  9. Lateral flow assays

    NARCIS (Netherlands)

    Posthuma-Trumpie, G.A.; Amerongen, van A.

    2012-01-01

    A simple version of immunochemical-based methods is the Lateral Flow Assay (LFA). It is a dry chemistry technique (reagents are included); the fluid from the sample runs through a porous membrane (often nitrocellulose) by capillary force. Typically the membrane is cut as a strip of 0.5*5 cm. In most

  10. Microchemiluminescent assay system

    Energy Technology Data Exchange (ETDEWEB)

    Kiel, J.L.

    1986-04-09

    The patent concerns a microchemiluminescent assay system, which can be used to detect ionizing radiation, heat or specific substances. The method involves the use of a complex formed from serum albumin and a luminescer which, in the presence of ionizing radiation (heat, or a specific analyte), will emit light in an amount proportional to the amount of radiation, etc. (U.K.).

  11. (MTT) dye reduction assay.

    African Journals Online (AJOL)

    to inhibit proliferation of HeLa cells was determined using the 3443- dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) dye reduction assay. Extracts from roots of Agathisanthemum bojeri, Synaptolepis kirkii and Zanha africana and the leaf extract of Physalis peruviana at a concentration of 10 pg/ml inhibited cell ...

  12. Hyaluronic Acid Assays

    DEFF Research Database (Denmark)

    Itenov, Theis S; Kirkby, Nikolai S; Bestle, Morten H

    2015-01-01

    BACKGROUD: Hyaluronic acid (HA) is proposed as a marker of functional liver capacity. The aim of the present study was to compare a new turbidimetric assay for measuring HA with the current standard method. METHODS: HA was measured by a particle-enhanced turbidimetric immunoassay (PETIA) and enzyme...

  13. FLUIDICS DEVICE FOR ASSAY

    DEFF Research Database (Denmark)

    2007-01-01

    The present invention relates to a device for use in performing assays on standard laboratory solid supports whereon chemical entities are attached. The invention furthermore relates to the use of such a device and a kit comprising such a device. The device according to the present invention is a...

  14. Radioreceptor assay for oxyphenonium

    International Nuclear Information System (INIS)

    Ensing, K.; Zeeuw, R.A. de

    1984-01-01

    The development of a radioreceptor assay for the quaternary anticholinergic drug, oxyphenonium, in plasma is reported. It is based on competition between this drug and 3 H-dexetimide for binding to muscarinic receptors. After ion pair extraction and reextraction, the drug can be determined in plasma at concentrations down to a value of 100 pg/ml. This permits pharmacokinetic studies to be made after inhalation of oxyphenonium. (author)

  15. Dual isotope assays

    International Nuclear Information System (INIS)

    Smith, G.F.W.; Stevens, R.A.J.; Jacoby, B.

    1980-01-01

    Dual isotope assays for thyroid function are performed by carrying out a radio-immunoassay for two of thyroxine (T4), tri-iodothyronine (T3), thyroid stimulating hormone (TSH), and thyroxine binding globulin (TBG), by a method wherein a version of one of the thyroid components, preferably T4 or T3 is labelled with Selenium-75 and the version of the other thyroid component is labelled with a different radionuclide, preferably Iodine-125. (author)

  16. A Quantitative Fluorescence-Based Lipase Assay

    Directory of Open Access Journals (Sweden)

    Giovanna Lomolino

    2012-01-01

    Full Text Available An easy and fast gel diffusion assay for detecting and monitoring lipase activity by quantification of fluorescein is described. By measuring the intensity of fluorescein, it is possible to obtain a calibration curve with a regression coefficient better than by using the radius of fluorescent haloes. Through the quantification of fluorescence intensity of fluorescein released after the hydrolysis of a fluorescent ester, fluorescein dibutyrate, used as substrate in agar plates, commercial and skimmed milk lipase activity were studied. Moreover, with this method, lipase activity can be monitored in reaction medium that contains compounds which are affected by turbidity or cause measurement interference for UV-spectrophotometer and fluorimeter. In this experiment, boiled skimmed milk was dispersed in the agar gel with fluorescein dibutyrate, and it was used as a reaction medium to mimic natural conditions. The development of such an assay has a potential for applications in industries ranging from pharmaceuticals to food production and monitoring.

  17. Radiorespirometic assay device

    International Nuclear Information System (INIS)

    Levin, G.V.; Straat, P.A.

    1981-01-01

    A radiorespirometic assay device is described in which the presence of microorganisms in a sample is determined by placing the sample in contact with a metabolisable radioactive labelled substrate, collecting any gas evolved, exposing a photosensitive material to the gas and determining if a spot is produced on the material. A spot indicates the presence of radioactivity showing that the substrate has been metabolized by a microorganism. Bacteria may be detected in body fluids, hospital operating rooms, water, food, cosmetics and drugs. (U.K.)

  18. Radon assay for SNO+

    Energy Technology Data Exchange (ETDEWEB)

    Rumleskie, Janet [Laurentian University, Greater Sudbury, Ontario (Canada)

    2015-12-31

    The SNO+ experiment will study neutrinos while located 6,800 feet below the surface of the earth at SNOLAB. Though shielded from surface backgrounds, emanation of radon radioisotopes from the surrounding rock leads to back-grounds. The characteristic decay of radon and its daughters allows for an alpha detection technique to count the amount of Rn-222 atoms collected. Traps can collect Rn-222 from various positions and materials, including an assay skid that will collect Rn-222 from the organic liquid scintillator used to detect interactions within SNO+.

  19. RAS - Screens & Assays - Drug Discovery

    Science.gov (United States)

    The RAS Drug Discovery group aims to develop assays that will reveal aspects of RAS biology upon which cancer cells depend. Successful assay formats are made available for high-throughput screening programs to yield potentially effective drug compounds.

  20. Improving shuffler assay accuracy

    International Nuclear Information System (INIS)

    Rinard, P.M.

    1995-01-01

    Drums of uranium waste should be disposed of in an economical and environmentally sound manner. The most accurate possible assays of the uranium masses in the drums are required for proper disposal. The accuracies of assays from a shuffler are affected by the type of matrix material in the drums. Non-hydrogenous matrices have little effect on neutron transport and accuracies are very good. If self-shielding is known to be a minor problem, good accuracies are also obtained with hydrogenous matrices when a polyethylene sleeve is placed around the drums. But for those cases where self-shielding may be a problem, matrices are hydrogenous, and uranium distributions are non-uniform throughout the drums, the accuracies are degraded. They can be greatly improved by determining the distributions of the uranium and then applying correction factors based on the distributions. This paper describes a technique for determining uranium distributions by using the neutron count rates in detector banks around the waste drum and solving a set of overdetermined linear equations. Other approaches were studied to determine the distributions and are described briefly. Implementation of this correction is anticipated on an existing shuffler next year

  1. Competitive protein binding assay

    International Nuclear Information System (INIS)

    Kaneko, Toshio; Oka, Hiroshi

    1975-01-01

    The measurement of cyclic GMP (cGMP) by competitive protein binding assay was described and discussed. The principle of binding assay was represented briefly. Procedures of our method by binding protein consisted of preparation of cGMP binding protein, selection of 3 H-cyclic GMP on market, and measurement procedures. In our method, binding protein was isolated from the chrysalis of silk worm. This method was discussed from the points of incubation medium, specificity of binding protein, the separation of bound cGMP from free cGMP, and treatment of tissue from which cGMP was extracted. cGMP existing in the tissue was only one tenth or one scores of cGMP, and in addition, cGMP competed with cGMP in binding with binding protein. Therefore, Murad's technique was applied to the isolation of cGMP. This method provided the measurement with sufficient accuracy; the contamination by cAMP was within several per cent. (Kanao, N.)

  2. Radioreceptor assay analysis of tamsulosin and terazosin pharmacokinetics

    NARCIS (Netherlands)

    Taguchi, K.; Schäfers, R. F.; Michel, M. C.

    1998-01-01

    AIMS: A radioreceptor assay has been developed for alpha1-adrenoceptor subtypes and applied to a pharmacokinetic analysis of tamsulosin and terazosin. METHODS: Young, male, healthy volunteers received 0.4 mg tamsulosin (as Omnic modified release capsules) or 5 mg terazosin (as Flotrin tablets) in a

  3. Toxics Release Inventory (TRI)

    Data.gov (United States)

    U.S. Environmental Protection Agency — The Toxics Release Inventory (TRI) is a dataset compiled by the U.S. Environmental Protection Agency (EPA). It contains information on the release and waste...

  4. Fluorescence imaging of glutamate release in neurons

    International Nuclear Information System (INIS)

    Wang, Ziqiang; Yeung, Edward S.

    1999-01-01

    A noninvasive detection scheme based on glutamate dehydrogenase (GDH) enzymatic assay combined with microscopy was developed to measure the glutamate release in cultured cells from the central nervous system (CNS). The enzyme reaction is very specific and sensitive. The detection limit with charge-coupled device (CCD) imaging is down to μM levels of glutamate with reasonable response time (∼30 s). The standard glutamate test shows a linear response over 3 orders of magnitude, from μM to 0.1 mM range. The in vitro monitoring of glutamate release from cultured neuron cells demonstrated excellent spatial and temporal resolution. (c) 1999 Society for Applied Spectroscopy

  5. An acoustic prion assay

    Directory of Open Access Journals (Sweden)

    Gordon Hayward

    2016-12-01

    Full Text Available An acoustic prion assay has been demonstrated for sheep brain samples. Only five false positives and no false negatives were observed in a test of 45 positive and 45 negative samples. The acoustic prion sensor was constructed using a thickness shear mode quartz resonator coated with a covalently bound recombinant prion protein. The characteristic indicator of a scrapie infected sheep brain sample was an observed shoulder in the frequency decrease in response to a sample.The response of the sensor aligns with a conformational shift in the surface protein and with the propagation mechanism of the disease. This alignment is evident in the response timing and shape, dependence on concentration, cross species behaviour and impact of blood plasma. This alignment is far from sufficient to prove the mechanism of the sensor but it does offer the possibility of a rapid and inexpensive additional tool to explore prion disease. Keywords: Prions, Thickness shear mode quartz sensor

  6. Assay of oestrogen

    International Nuclear Information System (INIS)

    Edwards, J.C.

    1981-01-01

    A particular problem with the direct radioimmunoassay of unconjugated oestriol in pregnancy is caused by the increased amount of steroid-binding proteins present in pregnancy serum and plasma. The steroid-binding proteins react with oestriol and 125 I-labelled oestriol during the assay procedure and the steroid-protein bound 125 I-labelled oestriol is precipitated along with the antibody-bound 125 I-labelled oestriol by the ammonium sulphate solution separation system. A novel method is described whereby progesterone (1-20 μg/ml) is used to block the action of steroid-binding proteins in pregnancy serum and plasma samples, thus minimizing interference in a direct radioimmunoassay for unconjugated oestriol using a specific anti-oestriol serum. (U.K.)

  7. An epidermal equivalent assay for identification and ranking potency of contact sensitizers

    NARCIS (Netherlands)

    Gibbs, S.; Corsini, E.; Spiekstra, S.W.; Galbiati, V.; Fuchs, H.W.; Degeorge, G.; Troese, M.; Hayden, P.; Deng, W.; Roggen, E.

    2013-01-01

    The purpose of this study was to explore the possibility of combining the epidermal equivalent (EE) potency assay with the assay which assesses release of interleukin-18 (IL-18) to provide a single test for identification and classification of skin sensitizing chemicals, including chemicals of low

  8. Influence of Seasonality and Circulating Cytokines on Serial QuantiFERON Discordances

    Directory of Open Access Journals (Sweden)

    Marsha L. Griffin

    2018-01-01

    Full Text Available Objectives. An 18-month prospective study serially tested healthcare workers (HCWs for tuberculosis infection (TBI and reported discordant QuantiFERON Gold In-Tube® (QFT results in some participants. The purpose of the current study was to investigate whether the interferon-gamma (IFN-γ measured by QFT in discordant individuals could be influenced by other circulating cytokines that vary seasonally at the time of phlebotomy. Methods. The CDC funded TBESC Task Order 18 (TO18 project to assess the use of Interferon Gamma Release Assays (IGRAs, T-SPOT.TB® and QFT, compared to the tuberculin skin test (TST for the serial testing of TBI in HCW at 4 US sites. Unstimulated plasma from 9 discordant TO18 participants at 4 different time points from the Houston site was multiplexed to determine the association between circulating cytokines and antigen stimulated IFN-γ levels. Results. IL-12, IL-1β, IL-3, GCSF, and IL-7 were associated with the amount of IFN-γ measured in response to antigen stimulation. In addition to these cytokines, a significant relationship was found between a positive QFT result and the spring season. Conclusions. Allergens during the spring season can result in the upregulation of IL-1β and IL-3, and this upregulation was observed with the amount of IFN-γ measured in discordant results.

  9. [Diagnostic strategies in the Tuberculosis Clinic of the Hospital General La Raza National Medical Center].

    Science.gov (United States)

    Flores-Ibarra, Alberto Alejandro; Ochoa-Vázquez, María Dolores; Sánchez-Tec, Georgina Alejandra

    2016-01-01

    In order to diagnose TB infection, tuberculin skin test and interferon gamma release assay are available. The tuberculin test has a sensitivity of 99 % and a specificity of 95 %. For the detection of interferon gamma in blood there are currently two tests available: TBGold QuantiFERON-In-Tube (with a sensitivity of 0.70 and a specificity of 0.90), and T-SPOT-TB (sensitivity 0.90 and specificity 0.93). To diagnose the disease, a microscopy of direct smears for acid-fast bacilli is used if the physician is facing an extensive cavitary lung disease due to M. tuberculosis (this test has a high sensitivity: 80-90 %). The most common staining techniques used are Ziehl-Neelsen and Kinyoun, and the fluorescent technique, auramine-rhodamine. The culture is the gold standard and it has a sensitivity of 80 % and a specificity over 90 %, but the results take weeks. The nucleic acid amplification test has an overall sensitivity and specificity of 0.85 and 0.97, respectively. In the presence of a pleural effusion is necessary to perform a pleural biopsy for culture with a sensitivity of 85 % if it is percutaneous and 98 % if it was taken by thoracoscopy. The adenosine deaminase can be determined in pleural fluid with a sensitivity and specificity of 95 %.

  10. Quantitative assay for the measurement of immune responses directed against the human placenta

    Energy Technology Data Exchange (ETDEWEB)

    Davies, M; Sutcliffe, R G [Glasgow Univ. (UK)

    1982-02-12

    A quantitative in vitro immune assay based on the classical chromium release assay has been developed to detect immune responses directed against alien antigens expressed by the developing foetus and present on the maternal-facing surface of the human placenta. A plasma membrane fraction from the surface of the placenta was prepared and the vesicles thus formed were radiolabelled with /sup 51/Cr. The /sup 51/Cr-labelled vesicles, by various criteria, were found to be suitable for use as targets in a release assay. Further, by means of experimentally immunised animals, the target membranes were shown to be capable of detecting both cellular and humoral anti-placental activity.

  11. Large scientific releases

    International Nuclear Information System (INIS)

    Pongratz, M.B.

    1981-01-01

    The motivation for active experiments in space is considered, taking into account the use of active techniques to obtain a better understanding of the natural space environment, the utilization of the advantages of space as a laboratory to study fundamental plasma physics, and the employment of active techniques to determine the magnitude, degree, and consequences of artificial modification of the space environment. It is pointed out that mass-injection experiments in space plasmas began about twenty years ago with the Project Firefly releases. Attention is given to mass-release techniques and diagnostics, operational aspects of mass release active experiments, the active observation of mass release experiments, active perturbation mass release experiments, simulating an artificial modification of the space environment, and active experiments to study fundamental plasma physics

  12. Evaluating pleural ADA, ADA2, IFN-γ and IGRA for diagnosing tuberculous pleurisy.

    Science.gov (United States)

    Keng, Li-Ta; Shu, Chin-Chung; Chen, Jason Yao-Ping; Liang, Sheng-Kai; Lin, Ching-Kai; Chang, Lih-Yu; Chang, Chia-Hao; Wang, Jann-Yuan; Yu, Chong-Jen; Lee, Li-Na

    2013-10-01

    Conventional methods for diagnosing tuberculous pleurisy (TB pleurisy) are either invasive or have a long turn-around-time. Performances of pleural adenosine deaminase (ADA), ADA2, interferon-gamma (IFN-γ), and interferon-gamma release assays (IGRA) as diagnostic tools for TB pleurisy were evaluated. Eighty-eight patients with lymphocyte-predominant pleural exudates between June 2010 and March 2011, including 31 with clinically diagnosed TB pleurisy, were prospectively studied. Pleural ADA and ADA2 activity were measured by colorimetric method, IFN-γ levels by enzyme-linked immuno-sorbent assay, and IGRA by enzyme-linked immuno-spot (T-SPOT.TB) assay. Pleural ADA, ADA2, and IFN-γ levels, but not the proportion of positive T-SPOT.TB assay, were significantly higher in patients with TB pleurisy than in those without TB pleurisy. The area under the receiver-operating-characteristic (ROC) curve was 0.920, 0.893, 0.875, and 0.544 for IFN-γ, ADA2, ADA, and T-SPOT.TB assay, respectively. The combination of ADA ≥ 40 IU/L and IFN-γ ≥ 75 pg/mL yielded a specificity of 100%. Pleural ADA, ADA2 and IFN-γ, but not T-SPOT.TB assay, are all sensitive and specific for TB pleurisy. In patients with lymphocyte-predominant pleural exudates, ADA ≥ 40 IU/L and IFN-γ ≥ 75 pg/mL in pleural effusion imply a very high probability of TB pleurisy. Copyright © 2013 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

  13. The Identification of Interferon-gamma as a Key Supportive Factor for Retinal Differentiation of Murine Mesenchymal Stem Cells

    Czech Academy of Sciences Publication Activity Database

    Heřmánková, Barbora; Kössl, Jan; Javorková, Eliška; Boháčová, Pavla; Hájková, Michaela; Zajícová, Alena; Krulová, Magdaléna; Holáň, Vladimír

    2017-01-01

    Roč. 26, č. 19 (2017), s. 1399-1408 ISSN 1547-3287 R&D Projects: GA ČR(CZ) GA17-04800S; GA MŠk(CZ) ED1.1.00/02.0109; GA MŠk(CZ) LO1309 Institutional support: RVO:68378041 Keywords : mesenchymal stem cell * differentiation * retina Subject RIV: FF - HEENT, Dentistry OBOR OECD: Cell biology Impact factor: 3.562, year: 2016

  14. Interferon Gamma-Based Detection of Latent Tuberculosis Infection in the Border States of Nuevo Leon and Tamaulipas, Mexico.

    Science.gov (United States)

    Oren, Eyal; Alatorre-Izaguirre, Gabriela; Vargas-Villarreal, Javier; Moreno-Treviño, Maria Guadalupe; Garcialuna-Martinez, Javier; Gonzalez-Salazar, Francisco

    2015-01-01

    Nearly one-third of the world's population is infected with latent tuberculosis (LTBI). Tuberculosis (TB) rates in the border states are higher than national rates in both the US and Mexico, with the border accounting for 30% of total registered TB cases in both countries. However, LTBI rates in the general population in Mexican border states are unknown. In this region, LTBI is diagnosed using the tuberculin skin test (TST). New methods of detection more specific than TST have been developed, although there is currently no gold standard for LTBI detection. Our objective is to demonstrate utility of the Quantiferon TB gold In-Tube (QFT-GIT) test compared with the TST to detect LTBI among border populations. This is an observational, cross-sectional study carried out in border areas of the states of Nuevo Leon and Tamaulipas, Mexico. Participants (n = 210) provided a TST and blood sample for the QFT-GIT. Kappa coefficients assessed the agreement between TST and QFT-GIT. Participant characteristics were compared using Fisher exact tests. Thirty-eight percent of participants were diagnosed with LTBI by QFT-GIT. The proportion of LTBI detected using QFT-GIT was almost double [38% (79/210)] that found by TST [19% (39/210)] (P < 0.001). Concordance between TST and QFT-GIT was low (kappa = 0.37). We recommend further studies utilizing the QFT-GIT test to detect LTBI among border populations.

  15. Association of interferon-gamma and interleukin 10 genotypes and serum levels with partial clinical remission in type 1 diabetes

    DEFF Research Database (Denmark)

    Alizadeh, B Z; Hanifi-Moghaddam, P; Eerligh, P

    2006-01-01

    We studied whether serum interferon (IFN)-gamma or interleukin (IL)-10 levels and their corresponding functional polymorphic genotypes are associated with partial remission of type 1 diabetes (T1D). A multi-centre study was undertaken in patients with newly diagnosed T1D and matched controls. T1D...

  16. Adjuvant interferon gamma in patients with pulmonary atypical Mycobacteriosis: A randomized, double-blind, placebo-controlled study

    Directory of Open Access Journals (Sweden)

    Sánchez-de la Osa Reinaldo B

    2008-02-01

    Full Text Available Abstract Background High antibiotic resistance is described in atypical Mycobacteriosis, mainly by Mycobacterium avium complex (MAC. Methods A randomized, double-blind, placebo-controlled clinical trial was carried out in two hospitals to evaluate the effect of interferon (IFN gamma as immunoadjuvant to chemotherapy on patients with atypical mycobacteria lung disease. Patients received placebo or 1 × 106 IU recombinant human IFN gamma intramuscularly, daily for one month and then three times per week up to 6 months as adjuvant to daily oral azithromycin, ciprofloxacin, ethambutol and rifampin. Sputum samples collection for direct smear observation and culture as well as clinical and thorax radiography assessments were done during treatment and one year after. Cytokines and oxidative stress determinations were carried out in peripheral blood before and after treatment. Results Eighteen patients were included in the IFN group and 14 received placebo. Groups were homogeneous at entry; average age was 60 years, 75% men, 84% white; MAC infection prevailed (94%. At the end of treatment, 72% of patients treated with IFN gamma were evaluated as complete responders, but only 36% in the placebo group. The difference was maintained during follow-up. A more rapid complete response was obtained in the IFN group (5 months before, with a significantly earlier improvement in respiratory symptoms and pulmonary lesions reduction. Disease-related deaths were 35.7% of the patients in the placebo group and only 11.1% in the IFN group. Three patients in the IFN group normalized their globular sedimentation rate values. Although differences in bacteriology were not significant during the treatment period, some patients in the placebo group converted again to positive during follow-up. Significant increments in serum TGF-beta and advanced oxidation protein products were observed in the placebo group but not among IFN receiving patients. Treatments were well tolerated. Flu-like symptoms predominated in the IFN gamma group. No severe events were recorded. Conclusion These data suggest that IFN gamma is useful and well tolerated as adjuvant therapy in patients with pulmonary atypical Mycobacteriosis, predominantly MAC. Further wider clinical trials are encouraged. Trial registration Current Controlled Trials ISRCTN70900209.

  17. Elevated interferon-gamma in CNS inflammatory disease: a potential complication for bone marrow reconstitution in MS

    DEFF Research Database (Denmark)

    Hassan-Zahraee, M; Tran, E H; Bourbonnière, L

    2000-01-01

    but levels were higher in IFNgamma transgenics. BM transplantation into IFNgamma-deficient recipients also had a high failure rate. Transplants of BM from mice lacking expression of IFNgamma-receptor failed, whereas IFNgamma-deficient grafts survived, suggesting that IFNgamma response status of the graft can......Bone marrow transplantation (BMT) is increasingly used to treat Multiple Sclerosis (MS) a CNS inflammatory disease with elevated CNS and systemic IFNgamma levels. We wished to determine the effect of IFNgamma on BM graft survival in a transgenic mouse model for chronic MS. BM transplantation...... into transgenic mice which express elevated levels of IFNgamma in the CNS was unsuccessful. By contrast, there was 100% survival of even fully allogeneic, T-depleted transplants to transgenics that over express TNFalpha in the CNS, using the same MBP promoter. IFNgamma was detectable in spleen of irradiated mice...

  18. Staphylococcal enterotoxin-A directly stimulates signal transduction and interferon-gamma production in psoriatic T-cell lines

    DEFF Research Database (Denmark)

    Nielsen, M B; Odum, N; Gerwien, J

    1998-01-01

    class II. Here we address the question of whether SEA can directly activate psoriatic T cells in the absence of professional antigen-presenting cells. We show that SEA induces i) tyrosine phosphorylation of several proteins, ii) downregulation of the T-cell receptor (TCR), and iii) production......-mediated proliferation. In contrast, SEA with a mutation in the MHC class II alpha-binding site induces IFN-gamma and a qualitatively changed tyrosine phosphorylation profile. Both mutations delete the co-stimulatory effect on cytokine-mediated proliferation. This suggests that both MHC class II binding sites...

  19. Interferon gamma responses to proteome-determined specific recombinant proteins: Potential as diagnostic markers for ovine Johne's disease

    Science.gov (United States)

    Johne’s disease (JD), or paratuberculosis is a fatal chronic granulomatous enteritis of animals caused by infection with Mycobacterium avium subspecies paratuberculosis (Map). A long subclinical phase may ensue during which time the animal shows no signs of clinical disease. Diagnosis of JD is probl...

  20. Interferon Gamma and PSA-Restricted Expression of FAS Ligand: A Novel Gene Therapy Strategy for Prostate Cancer

    National Research Council Canada - National Science Library

    Hall, Simon

    2003-01-01

    Introduction: Preliminary studies pointed to the ability for IFN-gamma to enhance sensitivity and/or reverse resistance to Fas transactivation on prostate cancer cells and work during the past 2 years illustrated...

  1. Opposite role of interferon-gamma and interleukin-4 on the regulation of blood pressure in mice

    NARCIS (Netherlands)

    Heuven-Nolsen, van D.; Kimpe, de S.J.; Muis, T.; Ark, van I.; Savelkoul, H.F.J.; Beems, R.B.; Oosterhout, van A.J.M.; Nijkamp, F.P.

    1999-01-01

    There is growing evidence that T-lymphocyte dysfunction contributes to the development of hypertension. IL-4 and IFN- are important regulators of T-lymphocyte function. Therefore, we investigated the effect of neutralizing antibodies against IL-4 (-IL-4) and IFN- (-IFN-) on the development of

  2. Hypoxia upregulates Bcl-2 expression and suppresses interferon-gamma induced antiangiogenic activity in human tumor derived endothelial cells.

    LENUS (Irish Health Repository)

    Wang, Jiang Huai

    2012-02-03

    BACKGROUND: Hypoxia in solid tumors potentially stimulates angiogenesis by promoting vascular endothelial growth factor (VEGF) production and upregulating VEGF receptor expression. However, it is unknown whether hypoxia can modulate the effect of anti-angiogenic treatment on tumor-derived endothelium. METHODS: Human tumor-derived endothelial cells (HTDEC) were freshly isolated from surgically removed human colorectal tumors by collagenase\\/DNase digestion and Percol gradient sedimentation. Cell proliferation was assessed by measuring BrdU incorporation, and capillary tube formation was measured using Matrigel. Cell apoptosis was assessed by flow cytometry and ELISA, and Bcl-2 expression was detected by Western blot analysis. RESULTS: Under aerobic culture conditions (5% CO2 plus 21% O2) HTDEC expressed less Bcl-2 and were more susceptible to IFN-gamma-induced apoptosis with significant reductions in both cell proliferation and capillary tube formation, when compared with normal human macrovascular and microvascular EC. Following exposure of HTDEC to hypoxia (5% CO2 plus 2% O2), IFN-gamma-induced cell apoptosis, and antiangiogenic activity (i.e. an inhibition in cell proliferation and capillary tube formation) in HTDEC were markedly attenuated. This finding correlated with hypoxia-induced upregulation of Bcl-2 expression in HTDEC. CONCLUSIONS: These results indicate that hypoxia can protect HTDEC against IFN-gamma-mediated cell death and antiangiogenic activity, and suggest that improvement of tumor oxygenation may potentiate the efficacy of anti-cancer therapies specifically targeting the inhibition of tumor angiogenesis.

  3. Association study of interferon gamma (IFN-γ) +874T/A gene polymorphism in patients with paranoid schizophrenia.

    Science.gov (United States)

    Paul-Samojedny, Monika; Owczarek, Aleksander; Suchanek, Renata; Kowalczyk, Malgorzata; Fila-Danilow, Anna; Borkowska, Paulina; Kucia, Krzysztof; Kowalski, Jan

    2011-03-01

    Schizophrenia is a multifactorial disease with changes affecting the immune system. Dysregulation of the cytokine network in schizophrenia has been well documented. Such changes may occur due to disturbances in cytokine levels that are linked to polymorphisms of cytokine genes. However, research in the role of cytokine gene polymorphisms in schizophrenia has been surprisingly scanty. The aim of this study was to identify, in a case control study, whether polymorphism of IFN-γ gene is a risk factor for the development of paranoid schizophrenia. To the best of our knowledge, this is the first study that examines the association between the IFN-γ gene polymorphism and psychopathological symptoms in patients with paranoid schizophrenia. Polymorphism of IFN-γ (+874T/A, rs 62559044) in schizophrenic patients (n=179), as well as healthy individuals (n=196), both Polish residents, was genotyped using AS-PCR method. Of note, when analyzing the results, we took into consideration the gender of studied individuals. Surprisingly, a single-nucleotide polymorphism in the first intron of the IFN-γ gene was found to be associated with paranoid schizophrenia in males, but not in females. The presence of allele A at position +874 in the IFN-γ gene correlates with 1.66-fold higher risk of paranoid schizophrenia development in males. Differences in the genotypes may have an important role in determining the level of I gene transcription. Because other polymorphisms have been demonstrated to influence IFN-γ transcription, further analysis is necessary to clarify the role of this gene in the pathogenesis of paranoid schizophrenia.

  4. Association between milk antibody and interferon-gamma responses in cattle from Mycobacterium avium subsp. paratuberculosis infected herds

    DEFF Research Database (Denmark)

    Mikkelsen, Heidi; Jungersen, Gregers; Nielsen, Søren Saxmose

    2009-01-01

    Paratuberculosis is a chronic infection of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). It is possible to detect infection with paratuberculosis at different stages of disease by means of various diagnostic test strategies. The objective of the present study was to evalu......Paratuberculosis is a chronic infection of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). It is possible to detect infection with paratuberculosis at different stages of disease by means of various diagnostic test strategies. The objective of the present study...

  5. Genetic variants of interferon-gamma and its mRNA expression and inflammatory parameters in the pathogenesis of vitiligo.

    Science.gov (United States)

    Karam, Rehab A; Zidan, Haidy E; Khater, Mohamed H

    2017-08-01

    Although genetics plays an essential role in the pathogenesis of vitiligo, vitiligo pathogenesis is still unclear. Our aim was to investigate the role of IFN-γ expression and polymorphism in vitiligo susceptibility and whether intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor (TNF)-α, and TNF-β play a role in vitiligo pathogenesis as important inflammatory parameters. Eighty-five patients with vitiligo and 90 controls were investigated for IFN-γ gene expression by quantitative real-time PCR and genotyped for IFN-γ +874T/A (rs2430561) and IFN-γ +2109A/G (rs1861494) gene polymorphisms by sequence-specific primer (SSP)-PCR and PCR-restriction fragment length polymorphism (RFLP), respectively. Serum levels of inflammatory parameters were measured using ELISA. Frequencies of the +874 TT genotype and T allele were significantly higher in patients with active vitiligo than in stable patients (P = 0.01 and 0.03, respectively). Calculation of odds ratio suggested a 1.7-fold increased risk of vitiligo in individuals having the TA haplotype. We observed overexpression of IFN-γ mRNA with elevated serum levels of IFN-γ, ICAM-1, TNF-α, and TNF-β in patients with vitiligo when compared with the control group (P = 0.001, for all). In addition, these levels were elevated in patients with active vitiligo compared with stable patients with vitiligo (P = 0.008, 0.006, 0.01, 0.01, and 0.03, respectively), which suggests the involvement of these cytokines in disease activity. In conclusion, IFN-γ is a promising immunological marker in vitiligo pathogenesis.

  6. Interferon-gamma (IFN-γ-mediated retinal ganglion cell death in human tyrosinase T cell receptor transgenic mouse.

    Directory of Open Access Journals (Sweden)

    Shahid Husain

    Full Text Available We have recently demonstrated the characterization of human tyrosinase TCR bearing h3T-A2 transgenic mouse model, which exhibits spontaneous autoimmune vitiligo and retinal dysfunction. The purpose of current study was to determine the role of T cells and IFN-γ in retina dysfunction and retinal ganglion cell (RGC death using this model. RGC function was measured by pattern electroretinograms (ERGs in response to contrast reversal of patterned visual stimuli. RGCs were visualized by fluorogold retrograde-labeling. Expression of CD3, IFN-γ, GFAP, and caspases was measured by immunohistochemistry and Western blotting. All functional and structural changes were measured in 12-month-old h3T-A2 mice and compared with age-matched HLA-A2 wild-type mice. Both pattern-ERGs (42%, p = 0.03 and RGC numbers (37%, p = 0.0001 were reduced in h3T-A2 mice when compared with wild-type mice. The level of CD3 expression was increased in h3T-A2 mice (h3T-A2: 174 ± 27% vs. HLA-A2: 100%; p = 0.04. The levels of effector cytokine IFN-γ were also increased significantly in h3T-A2 mice (h3T-A2: 189 ± 11% vs. HLA-A2: 100%; p = 0.023. Both CD3 and IFN-γ immunostaining were increased in nerve fiber (NF and RGC layers of h3T-A2 mice. In addition, we have seen a robust increase in GFAP staining in h3T-A2 mice (mainly localized to NF layer, which was substantially reduced in IFN-γ ((-/- knockout h3T-A2 mice. We also have seen an up-regulation of caspase-3 and -9 in h3T-A2 mice. Based on our data we conclude that h3T-A2 transgenic mice exhibit visual defects that are mostly associated with the inner retinal layers and RGC function. This novel h3T-A2 transgenic mouse model provides opportunity to understand RGC pathology and test neuroprotective strategies to rescue RGCs.

  7. Interferon-Gamma and Interlukin-4 Patterns in BALB/c Mice Suffering From Cutaneous Leishmaniasis Treated With Cantharidin

    OpenAIRE

    Maroufi, Yahya; Ghaffarifar, Fatemeh; Dalimi, Abdolhosein; Sharifi, Zohreh

    2014-01-01

    Background: Cutaneous leishmaniasis is a health problem in the world. Lesions should be treated on cosmetically or functionally important sites, such as the face and hands. Cantharidin is a terpenoid compound produced naturally by beetles of Meloidae and Oedemeridae families. Objectives: The current study aimed to investigate the effect of cantharidin on Cutaneous Leishmaniasis (CL) lesions and IFN-γ and IL-4 patterns in infected BALB/c mice. Materials and Methods: Infected BALB/c mice were d...

  8. Novel high-affinity binders of human interferon gamma derived from albumin-binding domain of protein G

    Czech Academy of Sciences Publication Activity Database

    Ahmad, Jawid Nazir; Li, J.; Biedermannová, Lada; Kuchař, Milan; Šípová, Hana; Semerádtová, A.; Černý, Jiří; Petroková, Hana; Mikulecký, Pavel; Polínek, Jiří; Staněk, Ondřej; Vondrášek, Jiří; Homola, Jiří; Malý, J.; Osička, Radim; Šebo, Peter; Malý, Petr

    2012-01-01

    Roč. 80, č. 3 (2012), s. 774-789 ISSN 0887-3585 R&D Projects: GA AV ČR KAN200520702; GA ČR GAP305/10/2184 Institutional research plan: CEZ:AV0Z50200510; CEZ:AV0Z50520701; CEZ:AV0Z20670512 Keywords : recombinant ligand * protein scaffold * computational design Subject RIV: CE - Biochemistry Impact factor: 3.337, year: 2012

  9. Interferon-gamma improves impaired dentinogenic and immunosuppressive functions of irreversible pulpitis-derived human dental pulp stem cells

    OpenAIRE

    Soichiro Sonoda; Haruyoshi Yamaza; Lan Ma; Yosuke Tanaka; Erika Tomoda; Reona Aijima; Kazuaki Nonaka; Toshio Kukita; Songtao Shi; Fusanori Nishimura; Takayoshi Yamaza

    2016-01-01

    Clinically, irreversible pulpitis is treated by the complete removal of pulp tissue followed by replacement with artificial materials. There is considered to be a high potential for autologous transplantation of human dental pulp stem cells (DPSCs) in endodontic treatment. The usefulness of DPSCs isolated from healthy teeth is limited. However, DPSCs isolated from diseased teeth with irreversible pulpitis (IP-DPSCs) are considered to be suitable for dentin/pulp regeneration. In this study, we...

  10. Assaying Cellular Viability Using the Neutral Red Uptake Assay.

    Science.gov (United States)

    Ates, Gamze; Vanhaecke, Tamara; Rogiers, Vera; Rodrigues, Robim M

    2017-01-01

    The neutral red uptake assay is a cell viability assay that allows in vitro quantification of xenobiotic-induced cytotoxicity. The assay relies on the ability of living cells to incorporate and bind neutral red, a weak cationic dye, in lysosomes. As such, cytotoxicity is expressed as a concentration-dependent reduction of the uptake of neutral red after exposure to the xenobiotic under investigation. The neutral red uptake assay is mainly used for hazard assessment in in vitro toxicology applications. This method has also been introduced in regulatory recommendations as part of 3T3-NRU-phototoxicity-assay, which was regulatory accepted in all EU member states in 2000 and in the OECD member states in 2004 as a test guideline (TG 432). The present protocol describes the neutral red uptake assay using the human hepatoma cell line HepG2, which is often employed as an alternative in vitro model for human hepatocytes. As an example, the cytotoxicity of acetaminophen and acetyl salicylic acid is assessed.

  11. A laboratory-scale pretreatment and hydrolysis assay for determination of reactivity in cellulosic biomass feedstocks.

    Science.gov (United States)

    Wolfrum, Edward J; Ness, Ryan M; Nagle, Nicholas J; Peterson, Darren J; Scarlata, Christopher J

    2013-11-14

    The rapid determination of the release of structural sugars from biomass feedstocks is an important enabling technology for the development of cellulosic biofuels. An assay that is used to determine sugar release for large numbers of samples must be robust, rapid, and easy to perform, and must use modest amounts of the samples to be tested.In this work we present a laboratory-scale combined pretreatment and saccharification assay that can be used as a biomass feedstock screening tool. The assay uses a commercially available automated solvent extraction system for pretreatment followed by a small-scale enzymatic hydrolysis step. The assay allows multiple samples to be screened simultaneously, and uses only ~3 g of biomass per sample. If the composition of the biomass sample is known, the results of the assay can be expressed as reactivity (fraction of structural carbohydrate present in the biomass sample released as monomeric sugars). We first present pretreatment and enzymatic hydrolysis experiments on a set of representative biomass feedstock samples (corn stover, poplar, sorghum, switchgrass) in order to put the assay in context, and then show the results of the assay applied to approximately 150 different feedstock samples covering 5 different materials. From the compositional analysis data we identify a positive correlation between lignin and structural carbohydrates, and from the reactivity data we identify a negative correlation between both carbohydrate and lignin content and total reactivity. The negative correlation between lignin content and total reactivity suggests that lignin may interfere with sugar release, or that more mature samples (with higher structural sugars) may have more recalcitrant lignin. The assay presented in this work provides a robust and straightforward method to measure the sugar release after pretreatment and saccharification that can be used as a biomass feedstock screening tool. We demonstrated the utility of the assay by

  12. The fluorometric microculture cytotoxicity assay.

    Science.gov (United States)

    Lindhagen, Elin; Nygren, Peter; Larsson, Rolf

    2008-01-01

    The fluorometric microculture cytotoxicity assay (FMCA) is a nonclonogenic microplate-based cell viability assay used for measurement of the cytotoxic and/or cytostatic effect of different compounds in vitro. The assay is based on hydrolysis of the probe, fluorescein diacetate (FDA) by esterases in cells with intact plasma membranes. The assay is available as both a semiautomated 96-well plate setup and a 384-well plate version fully adaptable to robotics. Experimental plates are prepared with a small amount of drug solution and can be stored frozen. Cells are seeded on the plates and cell viability is evaluated after 72 h. The protocol described here is applicable both for cell lines and freshly prepared tumor cells from patients and is suitable both for screening in drug development and as a basis for a predictive test for individualization of anticancer drug therapy.

  13. Solution assay instrument operations manual

    International Nuclear Information System (INIS)

    Li, T.K.; Marks, T.; Parker, J.L.

    1983-09-01

    An at-line solution assay instrument (SAI) has been developed and installed in a plutonium purification and americium recovery process area in the Los Alamos Plutonium Processing Facility. The instrument was designed for accurate, timely, and simultaneous nondestructive analysis of plutonium and americium in process solutions that have a wide range of concentrations and americium/plutonium ratios and for routine operation by process technicians who lack instrumentation background. The SAI, based on transmission-corrected, high-resolution gamma-ray spectroscopy, has two measurement stations attached to a single multichannel analyzer/computer system. To ensure the quality of assay results, the SAI has an internal measurement control program, which requires daily and weekly check runs and monitors key aspects of all assay runs. For a 25-ml sample, the assay precision is 5 g/l within a 2000-s count time

  14. The 2017 Release Cloudy

    Science.gov (United States)

    Ferland, G. J.; Chatzikos, M.; Guzmán, F.; Lykins, M. L.; van Hoof, P. A. M.; Williams, R. J. R.; Abel, N. P.; Badnell, N. R.; Keenan, F. P.; Porter, R. L.; Stancil, P. C.

    2017-10-01

    We describe the 2017 release of the spectral synthesis code Cloudy, summarizing the many improvements to the scope and accuracy of the physics which have been made since the previous release. Exporting the atomic data into external data files has enabled many new large datasets to be incorporated into the code. The use of the complete datasets is not realistic for most calculations, so we describe the limited subset of data used by default, which predicts significantly more lines than the previous release of Cloudy. This version is nevertheless faster than the previous release, as a result of code optimizations. We give examples of the accuracy limits using small models, and the performance requirements of large complete models. We summarize several advances in the H- and He-like iso-electronic sequences and use our complete collisional-radiative models to establish the densities where the coronal and local thermodynamic equilibrium approximations work.

  15. EIA new releases

    International Nuclear Information System (INIS)

    1994-09-01

    This report is a compliation of news releases from the Energy Information Administration. The september-october report includes articles on energy conservation, energy consumption in commercial buildings, and a short term energy model for a personal computer

  16. Radioligand assay in reproductive biology

    International Nuclear Information System (INIS)

    Korenman, S.G.; Sherman, B.M.

    1975-01-01

    Radioligand assays have been developed for the principal reproductive steroids and peptide hormones. Specific binding reagents have included antibodies, plasma binders, and intracellular receptors. In each assay, problems of specificity, sensitivity, and nonspecific inhibitors were encountered. Many features of the endocrine physiology in childhood, during puberty, and in adulthood have been characterized. Hormonal evaluations of endocrine disorders of reproduction are characterized on the basis of their characteristic pathophysiologic alterations. (U.S.)

  17. Sellafield (release of radioactivity)

    Energy Technology Data Exchange (ETDEWEB)

    Cunningham, J; Goodlad, A; Morris, M

    1986-02-06

    A government statement is reported, about the release of plutonium nitrate at the Sellafield site of British Nuclear Fuels plc on 5 February 1986. Matters raised included: details of accident; personnel monitoring; whether radioactive material was released from the site; need for public acceptance of BNFL activities; whether plant should be closed; need to reduce level of radioactive effluent; number of incidents at the plant.

  18. 3H-histamine release from human leukocytes

    International Nuclear Information System (INIS)

    Stahl Skov, P.; Norn, S.; Weeke, B.

    1979-01-01

    A rapid, simple, and inexpensive method for large scale screening of patients suspected of type I allergy has been developed. The method is based on in vitro incorporation of 3 H-histamine in the leukocytes of the patient, whereafter release of labelled histamine is measured after provocation of the cells with the suspected allergen. The new method was compared with the conventional basophil histamine release technique by in vitro provocation of six asthmatic patients under suspicion of type I allergy against animal dander, house dust, and mite, and an almost identical release of histamine was observed in both assays. (author)

  19. Correlation between the genotoxicity endpoints measured by two different genotoxicity assays: comet assay and CBMN assay

    Directory of Open Access Journals (Sweden)

    Carina Ladeira

    2015-06-01

    The results concerning of positive findings by micronuclei and non significant ones by comet assay, are corroborated by Deng et al. (2005 study performed in workers occupationally exposed to methotrexate, also a cytostatic drug. According to Cavallo et al. (2009, the comet assay seems to be more suitable for the prompt evaluation of the genotoxic effects, for instance, of polycyclic aromatic hydrocarbons mixtures containing volatile substances, whereas the micronucleus test seems more appropriate to evaluate the effects of exposure to antineoplastic agents. However, there are studies that observed an increase in both the comet assay and the micronucleus test in nurses handling antineoplastic drugs, although statistical significance was only seen in the comet assay, quite the opposite of our results (Maluf & Erdtmann, 2000; Laffon et al. 2005.

  20. ATP Release Channels

    Directory of Open Access Journals (Sweden)

    Akiyuki Taruno

    2018-03-01

    Full Text Available Adenosine triphosphate (ATP has been well established as an important extracellular ligand of autocrine signaling, intercellular communication, and neurotransmission with numerous physiological and pathophysiological roles. In addition to the classical exocytosis, non-vesicular mechanisms of cellular ATP release have been demonstrated in many cell types. Although large and negatively charged ATP molecules cannot diffuse across the lipid bilayer of the plasma membrane, conductive ATP release from the cytosol into the extracellular space is possible through ATP-permeable channels. Such channels must possess two minimum qualifications for ATP permeation: anion permeability and a large ion-conducting pore. Currently, five groups of channels are acknowledged as ATP-release channels: connexin hemichannels, pannexin 1, calcium homeostasis modulator 1 (CALHM1, volume-regulated anion channels (VRACs, also known as volume-sensitive outwardly rectifying (VSOR anion channels, and maxi-anion channels (MACs. Recently, major breakthroughs have been made in the field by molecular identification of CALHM1 as the action potential-dependent ATP-release channel in taste bud cells, LRRC8s as components of VRACs, and SLCO2A1 as a core subunit of MACs. Here, the function and physiological roles of these five groups of ATP-release channels are summarized, along with a discussion on the future implications of understanding these channels.

  1. Evaluation of tetrazolium-based semiautomatic colorimetric assay for measurement of human antitumor cytotoxicity

    International Nuclear Information System (INIS)

    Heo, D.S.; Park, J.G.; Hata, K.; Day, R.; Herberman, R.B.; Whiteside, T.L.

    1990-01-01

    A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)-based colorimetric assay was developed and compared with 51Cr release from different adherent tumor cell targets (human squamous cell carcinoma lines of the head and neck established in our laboratory, melanoma, and colorectal carcinoma) using 5-7-day human lymphokine-activated killer cells and monocyte-depleted peripheral blood lymphocytes as effectors. With adherent tumor cell targets, MTT colorimetry was more sensitive than the 51Cr release assay in measuring the antitumor activity of effectors: median, 4385 (range, 988-8144) versus median, 1061 (range, 582-7294) lytic units (the number of effector cells required to lyse 20% of 5 x 10(3) targets)/10(7) effectors (P less than 0.01). Background effects (without effector cells) were comparable in 4-h assays (9% versus 10%) between MTT colorimetry and 51Cr release. In 24-h assays, MTT colorimetry showed higher antitumor activity (70-100% versus 40-60% lysis at 1:1 effector:target cell ratio) but lower background effects (6% versus 38%) than 51Cr release assay. Thus, MTT colorimetry was more sensitive, did not use radiolabeled targets, required fewer effector cells, and was easier, less expensive, and better adaptable to serial monitoring of effector cell function in cancer patients. This colorimetric assay is especially well suited to adherent tumor cell targets. The use of adherent tumor cell monolayers, as opposed to trypsinized single cell suspensions, provides an opportunity to measure interactions of effector cells with enzymatically unaltered solid tumor targets. Because of the greater sensitivity of the colorimetric assay, the transformation of MTT data into lytic units, as commonly used for 51Cr release assays, required an adjustment to avoid the extrapolation based on the exponential fit equation

  2. Histamine release from cord blood basophils

    DEFF Research Database (Denmark)

    Nielsen, Bent Windelborg; Damsgaard, Tine Engberg; Herlin, Troels

    1990-01-01

    The histamine release (HR) after challenge with anti-IgE, concanavalin A, N-formyl-met-leu-phe and the calcium ionophore A23187 from 97 cord blood samples was determined by a microfiber-based assay. Maximum HR with anti-IgE showed great inter-individual variation (median: 20.5; range: 1-104 ng...... of less than 0.5 IU/ml, although sensitivity to anti-IgE was universally increased. Preincubation with pharmacologic agents modulating the IgE-mediated HR produced effects generally similar to previous findings in adult blood. However, the effects of inhibiting the cyclooxygenase pathway in cord blood...

  3. A simple and novel modification of comet assay for determination of bacteriophage mediated bacterial cell lysis.

    Science.gov (United States)

    Khairnar, Krishna; Sanmukh, Swapnil; Chandekar, Rajshree; Paunikar, Waman

    2014-07-01

    The comet assay is the widely used method for in vitro toxicity testing which is also an alternative to the use of animal models for in vivo testing. Since, its inception in 1984 by Ostling and Johansson, it is being modified frequently for a wide range of application. In spite of its wide applicability, unfortunately there is no report of its application in bacteriophages research. In this study, a novel application of comet assay for the detection of bacteriophage mediated bacterial cell lysis was described. The conventional methods in bacteriophage research for studying bacterial lysis by bacteriophages are plaque assay method. It is time consuming, laborious and costly. The lytic activity of bacteriophage devours the bacterial cell which results in the release of bacterial genomic material that gets detected by ethidium bromide staining method by the comet assay protocol. The objective of this study was to compare efficacy of comet assay with different assay used to study phage mediated bacterial lysis. The assay was performed on culture isolates (N=80 studies), modified comet assay appear to have relatively higher sensitivity and specificity than other assay. The results of the study showed that the application of comet assay can be an economical, time saving and less laborious alternative to conventional plaque assay for the detection of bacteriophage mediated bacterial cell lysis. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. RAVEN Beta Release

    International Nuclear Information System (INIS)

    Rabiti, Cristian; Alfonsi, Andrea; Cogliati, Joshua Joseph; Mandelli, Diego; Kinoshita, Robert Arthur; Wang, Congjian; Maljovec, Daniel Patrick; Talbot, Paul William

    2016-01-01

    This documents the release of the Risk Analysis Virtual Environment (RAVEN) code. A description of the RAVEN code is provided, and discussion of the release process for the M2LW-16IN0704045 milestone. The RAVEN code is a generic software framework to perform parametric and probabilistic analysis based on the response of complex system codes. RAVEN is capable of investigating the system response as well as the input space using Monte Carlo, Grid, or Latin Hyper Cube sampling schemes, but its strength is focused toward system feature discovery, such as limit surfaces, separating regions of the input space leading to system failure, using dynamic supervised learning techniques. RAVEN has now increased in maturity enough for the Beta 1.0 release.

  5. RAVEN Beta Release

    Energy Technology Data Exchange (ETDEWEB)

    Rabiti, Cristian [Idaho National Lab. (INL), Idaho Falls, ID (United States); Alfonsi, Andrea [Idaho National Lab. (INL), Idaho Falls, ID (United States); Cogliati, Joshua Joseph [Idaho National Lab. (INL), Idaho Falls, ID (United States); Mandelli, Diego [Idaho National Lab. (INL), Idaho Falls, ID (United States); Kinoshita, Robert Arthur [Idaho National Lab. (INL), Idaho Falls, ID (United States); Wang, Congjian [Idaho National Lab. (INL), Idaho Falls, ID (United States); Maljovec, Daniel Patrick [Idaho National Lab. (INL), Idaho Falls, ID (United States); Talbot, Paul William [Idaho National Lab. (INL), Idaho Falls, ID (United States)

    2016-02-01

    This documents the release of the Risk Analysis Virtual Environment (RAVEN) code. A description of the RAVEN code is provided, and discussion of the release process for the M2LW-16IN0704045 milestone. The RAVEN code is a generic software framework to perform parametric and probabilistic analysis based on the response of complex system codes. RAVEN is capable of investigating the system response as well as the input space using Monte Carlo, Grid, or Latin Hyper Cube sampling schemes, but its strength is focused toward system feature discovery, such as limit surfaces, separating regions of the input space leading to system failure, using dynamic supervised learning techniques. RAVEN has now increased in maturity enough for the Beta 1.0 release.

  6. Barcoded microchips for biomolecular assays.

    Science.gov (United States)

    Zhang, Yi; Sun, Jiashu; Zou, Yu; Chen, Wenwen; Zhang, Wei; Xi, Jianzhong Jeff; Jiang, Xingyu

    2015-01-20

    Multiplexed assay of analytes is of great importance for clinical diagnostics and other analytical applications. Barcode-based bioassays with the ability to encode and decode may realize this goal in a straightforward and consistent manner. We present here a microfluidic barcoded chip containing several sets of microchannels with different widths, imitating the commonly used barcode. A single barcoded microchip can carry out tens of individual protein/nucleic acid assays (encode) and immediately yield all assay results by a portable barcode reader or a smartphone (decode). The applicability of a barcoded microchip is demonstrated by human immunodeficiency virus (HIV) immunoassays for simultaneous detection of three targets (anti-gp41 antibody, anti-gp120 antibody, and anti-gp36 antibody) from six human serum samples. We can also determine seven pathogen-specific oligonucleotides by a single chip containing both positive and negative controls.

  7. Evolving BioAssay Ontology (BAO): modularization, integration and applications.

    Science.gov (United States)

    Abeyruwan, Saminda; Vempati, Uma D; Küçük-McGinty, Hande; Visser, Ubbo; Koleti, Amar; Mir, Ahsan; Sakurai, Kunie; Chung, Caty; Bittker, Joshua A; Clemons, Paul A; Brudz, Steve; Siripala, Anosha; Morales, Arturo J; Romacker, Martin; Twomey, David; Bureeva, Svetlana; Lemmon, Vance; Schürer, Stephan C

    2014-01-01

    The lack of established standards to describe and annotate biological assays and screening outcomes in the domain of drug and chemical probe discovery is a severe limitation to utilize public and proprietary drug screening data to their maximum potential. We have created the BioAssay Ontology (BAO) project (http://bioassayontology.org) to develop common reference metadata terms and definitions required for describing relevant information of low-and high-throughput drug and probe screening assays and results. The main objectives of BAO are to enable effective integration, aggregation, retrieval, and analyses of drug screening data. Since we first released BAO on the BioPortal in 2010 we have considerably expanded and enhanced BAO and we have applied the ontology in several internal and external collaborative projects, for example the BioAssay Research Database (BARD). We describe the evolution of BAO with a design that enables modeling complex assays including profile and panel assays such as those in the Library of Integrated Network-based Cellular Signatures (LINCS). One of the critical questions in evolving BAO is the following: how can we provide a way to efficiently reuse and share among various research projects specific parts of our ontologies without violating the integrity of the ontology and without creating redundancies. This paper provides a comprehensive answer to this question with a description of a methodology for ontology modularization using a layered architecture. Our modularization approach defines several distinct BAO components and separates internal from external modules and domain-level from structural components. This approach facilitates the generation/extraction of derived ontologies (or perspectives) that can suit particular use cases or software applications. We describe the evolution of BAO related to its formal structures, engineering approaches, and content to enable modeling of complex assays and integration with other ontologies and

  8. Hydraulic release oil tool

    International Nuclear Information System (INIS)

    Mims, M.G.; Mueller, M.D.; Ehlinger, J.C.

    1992-01-01

    This patent describes a hydraulic release tool. It comprises a setting assembly; a coupling member for coupling to drill string or petroleum production components, the coupling member being a plurality of sockets for receiving the dogs in the extended position and attaching the coupling member the setting assembly; whereby the setting assembly couples to the coupling member by engagement of the dogs in the sockets of releases from and disengages the coupling member in movement of the piston from its setting to its reposition in response to a pressure in the body in exceeding the predetermined pressure; and a relief port from outside the body into its bore and means to prevent communication between the relief port and the bore of the body axially of the piston when the piston is in the setting position and to establish such communication upon movement of the piston from the setting position to the release position and reduce the pressure in the body bore axially of the piston, whereby the reduction of the pressure signals that the tool has released the coupling member

  9. APASS Data Release 10

    Science.gov (United States)

    Henden, Arne A.; Levine, Stephen; Terrell, Dirk; Welch, Douglas L.; Munari, Ulisse; Kloppenborg, Brian K.

    2018-06-01

    The AAVSO Photometric All-Sky Survey (APASS) has been underway since 2010. This survey covers the entire sky from 7.5 knowledge of the optical train distortions. With these changes, DR10 includes many more stars than prior releases. We describe the survey, its remaining limitations, and prospects for the future, including a very-bright-star extension.

  10. Release the Prisoners Game

    Science.gov (United States)

    Van Hecke, Tanja

    2011-01-01

    This article presents the mathematical approach of the optimal strategy to win the "Release the prisoners" game and the integration of this analysis in a math class. Outline lesson plans at three different levels are given, where simulations are suggested as well as theoretical findings about the probability distribution function and its mean…

  11. Chromosome aberration assays in Allium

    Energy Technology Data Exchange (ETDEWEB)

    Grant, W.F.

    1982-01-01

    The common onion (Allium cepa) is an excellent plant for the assay of chromosome aberrations after chemical treatment. Other species of Allium (A. cepa var. proliferum, A. carinatum, A. fistulosum and A. sativum) have also been used but to a much lesser extent. Protocols have been given for using root tips from either bulbs or seeds of Allium cepa to study the cytological end-points, such as chromosome breaks and exchanges, which follow the testing of chemicals in somatic cells. It is considered that both mitotic and meiotic end-points should be used to a greater extent in assaying the cytogenetic effects of a chemical. From a literature survey, 148 chemicals are tabulated that have been assayed in 164 Allium tests for their clastogenic effect. Of the 164 assays which have been carried out, 75 are reported as giving a positive reaction, 49 positive and with a dose response, 1 positive and temperature-related, 9 borderline positive, and 30 negative; 76% of the chemicals gave a definite positive response. It is proposed that the Allium test be included among those tests routinely used for assessing chromosomal damage induced by chemicals.

  12. Assay of mast cell mediators

    DEFF Research Database (Denmark)

    Rådinger, Madeleine; Jensen, Bettina M; Swindle, Emily

    2015-01-01

    Mediator release from activated mast cells is a major initiator of the symptomology associated with allergic disorders such as anaphylaxis and asthma. Thus, methods to monitor the generation and release of such mediators have widespread applicability in studies designed to understand the processes...... regulating mast cell activation and for the identification of therapeutic approaches to block mast cell-driven disease. In this chapter, we discuss approaches used for the determination of mast cell degranulation, lipid-derived inflammatory mediator production, and cytokine/chemokine gene expression as well...

  13. [Managment of tuberculosis in an University of Campania].

    Science.gov (United States)

    Uccello, R; Monaco, M G L; Feola, D; Garzillo, E M; Muoio, M; Sannolo, N; Lamberti, M

    2012-01-01

    Tuberculosis (TBC) is an infectious disease with the highest mortality and morbidity by single pathogen, affecting about one third of worldwide population. Although Mantoux test is the most used, IGRA (Interferon-gamma Release Assays) tests seem to give good results for presumptive diagnosis of active or latent tuberculosis. From June 2011 to June 2012 we made about 1,000 visits for TBC prevention among the exposed to biological risks of our University. The management of suspected latent or active tuberculosis infection was carried out in collaboration with the pulmonologist, assessing the risk of contagion among exposed or affected operators. Health surveillance protocol and judgements of suitability for specific task were made not only in consideration of worker health, but also considerating the possible risk for patients, since this disease is a major problem for public health.

  14. Ocular Tuberculosis II: Diagnosis and Treatment

    Directory of Open Access Journals (Sweden)

    Sumru Önal

    2011-06-01

    Full Text Available Recent studies on the clinical importance of tuberculin skin tests (TST, interferon-gamma release assays (IGRAs, chest computed tomography (CT and polymerase chain reaction have provided a new approach to diagnosing ocular tuberculosis (TBC. However, both TST and IGRAs cannot distinguish between latent TBC infection and active disease. Another corroborative test in the diagnosis of intraocular TBC is the evaluation of the lungs by either chest radiography or CT. Direct evidence for the diagnosis can be obtained by examination of smears and staining for acid-fast organisms, cultures of intraocular tissue/fluid for Mycobacterium tuberculosis, and nucleic acid amplification analysis. In recent years, guidelines have been suggested for the diagnosis of intraocular TBC. The current treatment of intraocular TBC consists of use of four drugs (isoniazid, rifampicin, ethambutol and pyrazinamide taken for a long period of time (total 9-15 months. (Turk J Ophthalmol 2011; 41: 182-90

  15. Environmental mycobacteria – Future directions

    Directory of Open Access Journals (Sweden)

    D E Schraufnagel

    2015-01-01

    An interferon gamma release assay for infection with nontuberculous mycobacteria is desperately needed. Success with tuberculosis was attained by finding bacterial antigens associated with virulence and invasive disease. This may be difficult because the damage caused by environmental mycobacteria tends to be epithelial and subepithelial. The cavities may be bronchiectatic in origin and deeper invasion may result from host reactions. Serologic testing has not yielded important clinical help for diseases caused by either M. tuberculosis or the nontuberculous mycobacteria, possibly because good bacterial or host response proteins have not been identified. This could change with a metabolomics approach. Genomic studies that give incremental gains may set the stage for major breakthroughs, but require coordination with good phenotyping. Tools are in place to open an exciting new era for understanding and control of mycobacteria.

  16. IGRA-Based Screening for Latent Tuberculosis Infection in Persons Newly Incarcerated in New York City Jails.

    Science.gov (United States)

    Katyal, Monica; Leibowitz, Ruth; Venters, Homer

    2018-04-01

    In the United States, latent tuberculosis infection (LTBI) detection in correctional settings is a public health priority. Interferon gamma release assay (IGRA)-based LTBI screening was introduced in New York City jails in 2011 to 2012, replacing historically used tuberculin skin testing (TST), which was associated with substantial incomplete screening rates. This retrospective, cross-sectional study evaluated LTBI screening outcomes and correlates of positivity in 40,986 persons newly incarcerated in 2011 to 2013. Of 35,090 eligible patients tested (96.4%), final results were 6.3% positive, 93.4% negative, and 0.2% indeterminate. In multivariable regression modeling, sex, age, race/ethnicity, nativity, marital status, prior jail incarceration, and HIV status were correlated with positivity. IGRA-based screening yielded high screening and low indeterminate test rates and may be recommended in correctional and other settings where TST is currently used.

  17. Current management options for latent tuberculosis: a review

    Directory of Open Access Journals (Sweden)

    Norton BL

    2012-11-01

    Full Text Available Brianna L Norton, David P HollandDepartment of Medicine, Division of Infectious Diseases, Duke University Medical Center, Durham, NC, USAAbstract: Tuberculosis remains the world’s second leading infectious cause of death, with nearly one-third of the global population latently infected. Treatment of latent tuberculosis infection is a mainstay of tuberculosis-control efforts in low-to medium-incidence countries. Isoniazid monotherapy has been the standard of care for decades, but its utility is impaired by poor completion rates. However, new, shorter-course regimens using rifamycins improve completion rates and are cost-saving compared with standard isoniazid monotherapy. We review the currently available therapies for latent tuberculosis infection and their toxicities and include a brief economic comparison of the different regimens.Keywords: isoniazid, rifampin, rifapentine, tuberculin skin test, interferon-gamma release assay

  18. Development of a cell-based bioassay for phospholipase A2-triggered liposomal drug release

    DEFF Research Database (Denmark)

    Arouri, Ahmad; Trojnar, Jakub; Schmidt, Steffen

    2015-01-01

    models, the pattern of sPLA2-assisted drug release is unknown due to the lack of a suitable bio-relevant model. We report here on the development of a novel bioluminescence living-cell-based luciferase assay for the monitoring of sPLA2-triggered release of luciferin from liposomes. To this end, we...

  19. Automation of the dicentric chromosome assay and related assays

    International Nuclear Information System (INIS)

    Balajee, Adayabalam S.; Dainiak, Nicholas

    2016-01-01

    Dicentric Chromosome Assay (DCA) is considered to be the 'gold standard' for personalized dose assessment in humans after accidental or incidental radiation exposure. Although this technique is superior to other cytogenetic assays in terms of specificity and sensitivity, its potential application to radiation mass casualty scenarios is highly restricted because DCA is time consuming and labor intensive when performed manually. Therefore, it is imperative to develop high throughput automation techniques to make DCA suitable for radiological triage scenarios. At the Cytogenetic Biodosimetry Laboratory in Oak Ridge, efforts are underway to develop high throughput automation of DCA. Current status on development of various automated cytogenetic techniques in meeting the biodosimetry needs of radiological/nuclear incident(s) will be discussed

  20. Assay strategies and methods for phospholipases

    International Nuclear Information System (INIS)

    Reynolds, L.J.; Washburn, W.N.; Deems, R.A.; Dennis, E.A.

    1991-01-01

    Of the general considerations discussed, the two issues which are most important in choosing an assay are (1) what sensitivity is required to assay a particular enzyme and (2) whether the assay must be continuous. One can narrow the options further by considering substrate availability, enzyme specificity, assay convenience, or the presence of incompatible side reactions. In addition, the specific preference of a particular phospholipase for polar head group, micellar versus vesicular substrates, and anionic versus nonionic detergents may further restrict the options. Of the many assays described in this chapter, several have limited applicability or serious drawbacks and are not commonly employed. The most commonly used phospholipase assays are the radioactive TLC assay and the pH-stat assay. The TLC assay is probably the most accurate, sensitive assay available. These aspects often outweigh the disadvantages of being discontinuous, tedious, and expensive. The radioactive E. coli assay has become popular recently as an alternative to the TLC assay for the purification of the mammalian nonpancreatic phospholipases. The assay is less time consuming and less expensive than the TLC assay, but it is not appropriate when careful kinetics are required. Where less sensitivity is needed, or when a continuous assay is necessary, the pH-stat assay is often employed. With purified enzymes, when free thiol groups are not present, a spectrophotometric thiol assay can be used. This assay is ∼ as sensitive as the pH-stat assay but is more convenient and more reproducible, although the substrate is not available commercially. Despite the many assay choices available, the search continues for a convenient, generally applicable assay that is both sensitive and continuous

  1. Activated human neutrophils release hepatocyte growth factor/scatter factor.

    LENUS (Irish Health Repository)

    McCourt, M

    2012-02-03

    BACKGROUND: Hepatocyte growth factor or scatter factor (HGF\\/SF) is a pleiotropic cytokine that has potent angiogenic properties. We have previously demonstrated that neutrophils (PMN) are directly angiogenic by releasing vascular endothelial growth factor (VEGF). We hypothesized that the acute inflammatory response can stimulate PMN to release HGF. AIMS: To examine the effects of inflammatory mediators on PMN HGF release and the effect of recombinant human HGF (rhHGF) on PMN adhesion receptor expression and PMN VEGF release. METHODS: In the first experiment, PMN were isolated from healthy volunteers and stimulated with tumour necrosis factor-alpha (TNF-alpha), lipopolysaccharide (LPS), interleukin-8 (IL-8), and formyl methionyl-leucyl-phenylalanine (fMLP). Culture supernatants were assayed for HGF using ELISA. In the second experiment, PMN were lysed to measure total HGF release and HGF expression in the PMN was detected by Western immunoblotting. Finally, PMN were stimulated with rhHGF. PMN CD 11a, CD 11b, and CD 18 receptor expression and VEGF release was measured using flow cytometry and ELISA respectively. RESULTS: TNF-alpha, LPS and fMLP stimulation resulted in significantly increased release of PMN HGF (755+\\/-216, 484+\\/-221 and 565+\\/-278 pg\\/ml, respectively) compared to controls (118+\\/-42 pg\\/ml). IL-8 had no effect. Total HGF release following cell lysis and Western blot suggests that HGF is released from intracellular stores. Recombinant human HGF did not alter PMN adhesion receptor expression and had no effect on PMN VEGF release. CONCLUSIONS: This study demonstrates that pro-inflammatory mediators can stimulate HGF release from a PMN intracellular store and that activated PMN in addition to secreting VEGF have further angiogenic potential by releasing HGF.

  2. Myocardial myoglobin release after acute myocardial infarction

    Energy Technology Data Exchange (ETDEWEB)

    Robinson, P S; Saltissi, S; Coltart, D J; Croft, D N [Saint Thomas' Hospital, London (UK)

    1980-03-01

    The magnitude and time course of myoglobin release from the myocardium following infarction was assessed by radioimmunoassay. The assay showed acceptable precision over a working range from 50 to 750 ng cm/sup -3/, provided careful control of the assay temperature was maintained. The use of this radioimmunoassay as an early diagnostic test for infarction and as a potential measure of the extent of necrosis is considered and comparison made with the release of CK-MB, the myocardial specific isoenzyme of creatine kinase. Of the twenty patients studied with myocardial infarction, all had elevated levels of serum myoglobin including those admitted within 3 hours of the onset of pain. In contrast, CK-MB was not detected in the serum within 5 hours of the onset of pain. Peak serum levels of myoglobin (mean 852 +- 365 ng cm/sup -3/) and CK-MB (mean 71 +- 25 mIU cm/sup -3/) were detected at 8-16 hours and 20-24 hours respectively after the onset of pain. A comparison of peak serum levels of myoglobin and CK-MB showed a good correlation (r = 0.84).

  3. Stretch-induced Ca2+ independent ATP release in hippocampal astrocytes.

    Science.gov (United States)

    Xiong, Yingfei; Teng, Sasa; Zheng, Lianghong; Sun, Suhua; Li, Jie; Guo, Ning; Li, Mingli; Wang, Li; Zhu, Feipeng; Wang, Changhe; Rao, Zhiren; Zhou, Zhuan

    2018-02-28

    Similar to neurons, astrocytes actively participate in synaptic transmission via releasing gliotransmitters. The Ca 2+ -dependent release of gliotransmitters includes glutamate and ATP. Following an 'on-cell-like' mechanical stimulus to a single astrocyte, Ca 2+ independent single, large, non-quantal, ATP release occurs. Astrocytic ATP release is inhibited by either selective antagonist treatment or genetic knockdown of P2X7 receptor channels. Our work suggests that ATP can be released from astrocytes via two independent pathways in hippocampal astrocytes; in addition to the known Ca 2+ -dependent vesicular release, larger non-quantal ATP release depends on P2X7 channels following mechanical stretch. Astrocytic ATP release is essential for brain functions such as synaptic long-term potentiation for learning and memory. However, whether and how ATP is released via exocytosis remains hotly debated. All previous studies of non-vesicular ATP release have used indirect assays. By contrast, two recent studies report vesicular ATP release using more direct assays. In the present study, using patch clamped 'ATP-sniffer cells', we re-investigated astrocytic ATP release at single-vesicle resolution in hippocampal astrocytes. Following an 'on-cell-like' mechanical stimulus of a single astrocyte, a Ca 2+ independent single large non-quantal ATP release occurred, in contrast to the Ca 2+ -dependent multiple small quantal ATP release in a chromaffin cell. The mechanical stimulation-induced ATP release from an astrocyte was inhibited by either exposure to a selective antagonist or genetic knockdown of P2X7 receptor channels. Functional P2X7 channels were expressed in astrocytes in hippocampal brain slices. Thus, in addition to small quantal ATP release, larger non-quantal ATP release depends on P2X7 channels in astrocytes. © 2018 The Authors. The Journal of Physiology © 2018 The Physiological Society.

  4. Decontamination for free release

    Energy Technology Data Exchange (ETDEWEB)

    Simpson, K A; Elder, G R [Bradtec Ltd., Bristol (United Kingdom)

    1997-02-01

    Many countries are seeking to treat radioactive waste in ways which meet the local regulatory requirements, but yet are cost effective when all contributing factors are assessed. In some countries there are increasing amounts of waste, arising from nuclear plant decommissioning, which are categorized as low level waste: however with suitable treatment a large part of such wastes might become beyond regulatory control and be able to be released as non-radioactive. The benefits and disadvantages of additional treatment before disposal need to be considered. Several processes falling within the overall description of decontamination for free release have been developed and applied, and these are outlined. In one instance the process seeks to take advantage of techniques and equipment used for decontaminating water reactor circuits intermittently through reactor life. (author). 9 refs, 1 fig., 3 tabs.

  5. Atmospheric Release Advisory Capability

    International Nuclear Information System (INIS)

    Dickerson, M.H.; Gudiksen, P.H.; Sullivan, T.J.

    1983-02-01

    The Atmospheric Release Advisory Capability (ARAC) project is a Department of Energy (DOE) sponsored real-time emergency response service available for use by both federal and state agencies in case of a potential or actual atmospheric release of nuclear material. The project, initiated in 1972, is currently evolving from the research and development phase to full operation. Plans are underway to expand the existing capability to continuous operation by 1984 and to establish a National ARAC Center (NARAC) by 1988. This report describes the ARAC system, its utilization during the past two years, and plans for its expansion during the next five to six years. An integral part of this expansion is due to a very important and crucial effort sponsored by the Defense Nuclear Agency to extend the ARAC service to approximately 45 Department of Defense (DOD) sites throughout the continental US over the next three years

  6. Border cell release

    DEFF Research Database (Denmark)

    Mravec, Jozef

    2017-01-01

    Plant border cells are specialised cells derived from the root cap with roles in the biomechanics of root growth and in forming a barrier against pathogens. The mechanism of highly localised cell separation which is essential for their release to the environment is little understood. Here I present...... in situ analysis of Brachypodium distachyon, a model organism for grasses which possess type II primary cell walls poor in pectin content. Results suggest similarity in spatial dynamics of pectic homogalacturonan during dicot and monocot border cell release. Integration of observations from different...... species leads to the hypothesis that this process most likely does not involve degradation of cell wall material but rather employs unique cell wall structural and compositional means enabling both the rigidity of the root cap as well as detachability of given cells on its surface....

  7. Energy released in fission

    International Nuclear Information System (INIS)

    James, M.F.

    1969-05-01

    The effective energy released in and following the fission of U-235, Pu-239 and Pu-241 by thermal neutrons, and of U-238 by fission spectrum neutrons, is discussed. The recommended values are: U-235 ... 192.9 ± 0.5 MeV/fission; U-238 ... 193.9 ± 0.8 MeV/fission; Pu-239 ... 198.5 ± 0.8 MeV/fission; Pu-241 ... 200.3 ± 0.8 MeV/fission. These values include all contributions except from antineutrinos and very long-lived fission products. The detailed contributions are discussed, and inconsistencies in the experimental data are pointed out. In Appendix A, the contribution to the total useful energy release in a reactor from reactions other than fission are discussed briefly, and in Appendix B there is a discussion of the variations in effective energy from fission with incident neutron energy. (author)

  8. Slow-release fertilizer

    Science.gov (United States)

    Ming, Douglas W.; Golden, D. C.

    1992-10-01

    A synthetic apatite containing agronutrients and a method for making the apatite are disclosed. The apatite comprises crystalline calcium phosphate having agronutrients dispersed in the crystalline structure. The agronutrients can comprise potassium, magnesium, sulfur, iron, manganese, molybdenum, chlorine, boron, copper and zinc in amounts suited for plant growth. The apatite can optionally comprise a carbonate and/or silicon solubility control agent. The agronutrients are released slowly as the apatite dissolves.

  9. EIA new releases

    International Nuclear Information System (INIS)

    1994-12-01

    This report was prepared by the Energy Information Administration. It contains news releases on items of interest to the petroleum, coal, nuclear, electric and alternate fuels industries ranging from economic outlooks to environmental concerns. There is also a listing of reports by industry and an energy education resource listing containing sources for free or low-cost energy-related educational materials for educators and primary and secondary students

  10. Atmospheric release advisory capability

    International Nuclear Information System (INIS)

    Sullivan, T.J.

    1981-01-01

    The ARAC system (Atmospheric Release Advisory Capability) is described. The system is a collection of people, computers, computer models, topographic data and meteorological input data that together permits a calculation of, in a quasi-predictive sense, where effluent from an accident will migrate through the atmosphere, where it will be deposited on the ground, and what instantaneous and integrated dose an exposed individual would receive

  11. Slow-release fertilizer

    Science.gov (United States)

    Ming, Douglas W. (Inventor); Golden, Dadigamuwage C. (Inventor)

    1995-01-01

    A synthetic apatite containing agronutrients and a method for making the apatite are disclosed. The apatite comprises crystalline calcium phosphate having agronutrients dispersed in the crystalline structure. The agronutrients can comprise potassium, magnesium, sulfur, iron, manganese, molybdenum, chlorine, boron, copper and zinc in amounts suited for plant growth. The apatite can optionally comprise a carbonate and/or silicon solubility control agent. The agronutrients are released slowly as the apatite dissolves.

  12. Contact: Releasing the news

    Science.gov (United States)

    Pinotti, Roberto

    The problem of mass behavior after man's future contacts with other intelligences in the universe is not only a challenge for social scientists and political leaders all over the world, but also a cultural time bomb as well. In fact, since the impact of CETI (Contact with Extraterrestrial Intelligence) on human civilization, with its different cultures, might cause a serious socio-anthropological shock, a common and predetermined worldwide strategy is necessary in releasing the news after the contact, in order to keep possible manifestations of fear, panic and hysteria under control. An analysis of past studies in this field and of parallel historical situations as analogs suggests a definite "authority crisis" in the public as a direct consequence of an unexpected release of the news, involving a devastating "chain reaction" process (from both the psychological and sociological viewpoints) of anomie and maybe the collapse of today's society. The only way to prevent all this is to prepare the world's public opinion concerning contact before releasing the news, and to develop a long-term strategy through the combined efforts of scientists, political leaders, intelligence agencies and the mass media, in order to create the cultural conditions in which a confrontation with ETI won't affect mankind in a traumatic way. Definite roles and tasks in this multi-level model are suggested.

  13. Nondestructive assay of sale materials

    International Nuclear Information System (INIS)

    Rodenburg, W.W.; Fleissner, J.G.

    1981-01-01

    This paper covers three primary areas: (1) reasons for performing nondestructive assay on SALE materials; (2) techniques used; and (3) discussion of investigators' revised results. The study shows that nondestructive calorimetric assay of plutonium offers a viable alternative to traditional wet chemical techniques. For these samples, the precision ranged from 0.4 to 0.6% with biases less than 0.2%. Thus, for those materials where sampling errors are the predominant source of uncertainty, this technique can provide improved accuracy and precision while saving time and money as well as reducing the amount of liquid wastes to be handled. In addition, high resolution gamma-ray spectroscopy measurements of solids can provide isotopic analysis data in a cost effective and timely manner. The timeliness of the method can be especially useful to the plant operator for production control and quality control measurements

  14. Comet Assay in Cancer Chemoprevention.

    Science.gov (United States)

    Santoro, Raffaela; Ferraiuolo, Maria; Morgano, Gian Paolo; Muti, Paola; Strano, Sabrina

    2016-01-01

    The comet assay can be useful in monitoring DNA damage in single cells caused by exposure to genotoxic agents, such as those causing air, water, and soil pollution (e.g., pesticides, dioxins, electromagnetic fields) and chemo- and radiotherapy in cancer patients, or in the assessment of genoprotective effects of chemopreventive molecules. Therefore, it has particular importance in the fields of pharmacology and toxicology, and in both environmental and human biomonitoring. It allows the detection of single strand breaks as well as double-strand breaks and can be used in both normal and cancer cells. Here we describe the alkali method for comet assay, which allows to detect both single- and double-strand DNA breaks.

  15. Radioreceptor assay for somatomedin A

    Energy Technology Data Exchange (ETDEWEB)

    Takano, K [Tokyo Women' s Medical Coll. (Japan)

    1975-04-01

    Measurement method of somatomedian A by radioreceptor assay using the human placenta membrane was described and discussed. Binding rate of /sup 125/I-somatomedin A to its receptors was studied under various conditions of time and temperature of the incubation, and pH of the system. The influence of somatomedin A, porcine insulin, and porcine calcitonin, on /sup 125/I-somatomedin A bound receptors was studied, and these hormones showed the competitive binding to somatomedin A receptors in some level. The specificity, recovery rate, and clinical applications of somatomedin A were also discussed. Radioreceptor assay for somatomedine A provided easier, faster, and more accurate measurements than conventional bioassay. This technique would be very useful to study somatomedin A receptor and functions of insulin.

  16. Tratamento da tuberculose de infecção latente. As recomendações actuais

    Directory of Open Access Journals (Sweden)

    R. Duarte

    2010-09-01

    Full Text Available Resumo: O diagnóstico e tratamento da infecção latente por Mycobacterium tuberculosis reduz significativamente o risco de desenvolvimento de tuberculose activa e a transmissão da doença na comunidade. O rastreio da tuberculose infecção latente deve passar pela exclusão de doença activa (inquérito de sintomas e radiografia pulmonar e avaliação da resposta imunológica ao M. tuberculosis através dos testes actualmente ao dispor, como o teste tuberculínico e os testes IGRA (interferon-gamma release assay. A escolha do esquema de tratamento deve ter em linha de conta a eficácia, a adesão e os efeitos colaterais associados ao mesmo Este documento actualiza as recomendações sobre tratamento da tuberculose infecção latente. São apresentadas indicações sobre quem deve ser rastreado e revistos os esquemas de tratamento. Abstract: Diagnosis and treatment of latent infection with Mycobacterium tuberculosis (LTBI, significantly reduces the risk of developing active tuberculosis and the transmission of the disease in the community. LTBI screening must pass by the exclusion of active disease (symptoms enquiry and chest radiography and assessment of immune response to Mycobacterium tuberculosis testing with the tests currently available – tuberculin skin test and interferon-gamma release assay (IGRA. The choice of treatment must take into account the efficacy and side effects associated with the same. This document provides updated recommendations on latent tuberculosis infection treatment. Topics covered include whom to test for TB and reviewed LTBI treatment regimens. Palavras-chave: Tuberculose, tuberculose infeccao latente, tratamento, preventivo, Key words: Tuberculosis, latent tuberculosis infection, treatment, preventive

  17. Tratamento da tuberculose de infecção latente: As recomendações actuais Latent tuberculosis infection treatment: Current recommendations

    Directory of Open Access Journals (Sweden)

    R Duarte

    2010-09-01

    Full Text Available O diagnóstico e tratamento da infecção latente por Mycobacterium tuberculosis reduz significativamente o risco de desenvolvimento de tuberculose activa e a transmissão da doença na comunidade. O rastreio da tuberculose infecção latente deve passar pela exclusão de doença activa (inquérito de sintomas e radiografia pulmonar e avaliação da resposta imunológica ao M. tuberculosis através dos testes actualmente ao dispor, como o teste tuberculínico e os testes IGRA (interferon-gamma release assay. A escolha do esquema de tratamento deve ter em linha de conta a eficácia, a adesão e os efeitos colaterais associados ao mesmo Este documento actualiza as recomendações sobre tratamento da tuberculose infecção latente. São apresentadas indicações sobre quem deve ser rastreado e revistos os esquemas de tratamento.Diagnosis and treatment of latent infection with Mycobacterium tuberculosis (LTBI, significantly reduces the risk of developing active tuberculosis and the transmission of the disease in the community. LTBI screening must pass by the exclusion of active disease (symptoms enquiry and chest radiography and assessment of immune response to Mycobacterium tuberculosis testing with the tests currently available - tuberculin skin test and interferon-gamma release assay (IGRA. The choice of treatment must take into account the efficacy and side effects associated with the same. This document provides updated recommendations on latent tuberculosis infection treatment. Topics covered include whom to test for TB and reviewed LTBI treatment regimens.

  18. Assay of vitamin B12

    International Nuclear Information System (INIS)

    Tovey, K.C.; Carrick, D.T.

    1982-01-01

    A radioassay is described for vitamin B12 which involves denaturing serum protein binding proteins with alkali. In the denaturation step a dithiopolyol and cyanide are used and in the intrinsic factor assay step a vitamin B12 analogue such as cobinamide is used to bind with any remaining serum proteins. The invention also includes a kit in which the dithiopolyol is provided in admixture with the alkali. The dithiopolyol may be dithiothreitol or dithioerythritol. (author)

  19. Assay of ribulose bisphosphate carboxylase

    International Nuclear Information System (INIS)

    Pike, C.; Berry, J.

    1987-01-01

    Assays of ribulose bisphosphate carboxylase (rubisco) can be used to illustrate many properties of photosynthetic systems. Many different leaves have been assayed with this standard procedure. The tissue is ground with a mortar and pestle in extraction buffer. The supernatant after centrifugation is used as the source of enzyme. Buffer, RuBP, [ 14 C]-NaHCO 3 , and enzyme are combined in a scintillation vial; the reaction is run for 1 min at 30 0 . The acid-stable products are counted. Reproducibility in student experiments has been excellent. The assay data can be combined with analyses of leaf properties such as fresh and dry weight, chlorophyll and protein content, etc. Students have done projects such as the response of enzyme to temperature and to various inhibitors. They also report on the use of a transition state analog, carboxyarabinitol bisphosphate, to titrate the molar concentration of rubisco molecules (active sites) in an enzyme sample. Thus, using crude extracts the catalytic activity of a sample can be compared to the absolute quantity of enzyme or to the turnover number

  20. Protecting privacy in data release

    CERN Document Server

    Livraga, Giovanni

    2015-01-01

    This book presents a comprehensive approach to protecting sensitive information when large data collections are released by their owners. It addresses three key requirements of data privacy: the protection of data explicitly released, the protection of information not explicitly released but potentially vulnerable due to a release of other data, and the enforcement of owner-defined access restrictions to the released data. It is also the first book with a complete examination of how to enforce dynamic read and write access authorizations on released data, applicable to the emerging data outsou

  1. Triggered Release from Polymer Capsules

    Energy Technology Data Exchange (ETDEWEB)

    Esser-Kahn, Aaron P. [Univ. of Illinois, Urbana, IL (United States). Beckman Inst. for Advanced Science and Technology and Dept. of Chemistry; Odom, Susan A. [Univ. of Illinois, Urbana, IL (United States). Beckman Inst. for Advanced Science and Technology and Dept. of Chemistry; Sottos, Nancy R. [Univ. of Illinois, Urbana, IL (United States). Beckman Inst. for Advanced Science and Technology and Dept. of Materials Science and Engineering; White, Scott R. [Univ. of Illinois, Urbana, IL (United States). Beckman Inst. for Advanced Science and Technology and Dept. of Aerospace Engineering; Moore, Jeffrey S. [Univ. of Illinois, Urbana, IL (United States). Beckman Inst. for Advanced Science and Technology and Dept. of Chemistry

    2011-07-06

    Stimuli-responsive capsules are of interest in drug delivery, fragrance release, food preservation, and self-healing materials. Many methods are used to trigger the release of encapsulated contents. Here we highlight mechanisms for the controlled release of encapsulated cargo that utilize chemical reactions occurring in solid polymeric shell walls. Triggering mechanisms responsible for covalent bond cleavage that result in the release of capsule contents include chemical, biological, light, thermal, magnetic, and electrical stimuli. We present methods for encapsulation and release, triggering methods, and mechanisms and conclude with our opinions on interesting obstacles for chemically induced activation with relevance for controlled release.

  2. Quality Control Assays for Clinical-Grade Human Mesenchymal Stromal Cells: Validation Strategy.

    Science.gov (United States)

    Radrizzani, Marina; Soncin, Sabrina; Bolis, Sara; Lo Cicero, Viviana; Andriolo, Gabriella; Turchetto, Lucia

    2016-01-01

    The present chapter focuses on the validation of the following analytical methods for the control of mesenchymal stromal cells (MSC) for cell therapy clinical trials: Microbiological control for cellular product Endotoxin assay Mycoplasma assay Cell count and viability Immunophenotype Clonogenic potential (CFU-F assay) In our lab, these methods are in use for product release, process control or control of the biological starting materials. They are described in detail in the accompanying Chapter 19.For each method, validation goals and strategy are presented, and a detailed experimental scheme is proposed.

  3. A continuous spectrophotometric assay for monitoring adenosine 5'-monophosphate production.

    Science.gov (United States)

    First, Eric A

    2015-08-15

    A number of biologically important enzymes release adenosine 5'-monophosphate (AMP) as a product, including aminoacyl-tRNA synthetases, cyclic AMP (cAMP) phosphodiesterases, ubiquitin and ubiquitin-like ligases, DNA ligases, coenzyme A (CoA) ligases, polyA deadenylases, and ribonucleases. In contrast to the abundance of assays available for monitoring the conversion of adenosine 5'-triphosphate (ATP) to ADP, there are relatively few assays for monitoring the conversion of ATP (or cAMP) to AMP. In this article, we describe a homogeneous assay that continuously monitors the production of AMP. Specifically, we have coupled the conversion of AMP to inosine 5'-monophosphate (IMP) (by AMP deaminase) to the oxidation of IMP (by IMP dehydrogenase). This results in the reduction of oxidized nicotine adenine dinucleotide (NAD(+)) to reduced nicotine adenine dinucleotide (NADH), allowing AMP formation to be monitored by the change in the absorbance at 340 nm. Changes in AMP concentrations of 5 μM or more can be reliably detected. The ease of use and relatively low expense make the AMP assay suitable for both high-throughput screening and kinetic analyses. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Riola release report

    Energy Technology Data Exchange (ETDEWEB)

    Woodward, E.C.

    1983-08-04

    Eleven hours after execution of the Riola Event (at 0826 PDT on 25 September 1980) in hole U2eq of the Nevada Test Site (NTS), a release of radioactivity began. When the seepage stopped at about noon the following day, up to some 3200 Ci of activity had been dispersed by light variable winds. On 26 September, examination of the geophone records showed six hours of low-level, but fairly continuous, activity before the release. Electrical measurements indicated that most cables were still intact to a depth below the stemming platform. A survey of the ground zero area showed that the seepage came through cracks between the surface conductor and the pad, through cracks in the pad, and through a crack adjacent to the pad around the mousehole (a small hole adjacent to the emplacement hole). To preclude undue radiation exposure or injury from a surprise subsidence, safety measures were instituted. Tritium seepage was suffucient to postpone site activities until a box and pipeline were emplaced to contain and remove the gas. Radiation release modeling and calculations were generally consistent with observations. Plug-hole interaction calculations showed that the alluvium near the bottom of the plug may have been overstressed and that improvements in the design of the plug-medium interface can be made. Experimental studies verified that the surface appearance of the plug core was caused by erosion, but, assuming a normal strength for the plug material, that erosion alone could not account for the disappearance of such a large portion of the stemming platform. Samples from downhole plug experiments show that the plug may have been considerably weaker than had been indicted by quality assurance (QA) samples. 19 references, 32 figures, 10 tables.

  5. Riola release report

    International Nuclear Information System (INIS)

    Woodward, E.C.

    1983-01-01

    Eleven hours after execution of the Riola Event (at 0826 PDT on 25 September 1980) in hole U2eq of the Nevada Test Site (NTS), a release of radioactivity began. When the seepage stopped at about noon the following day, up to some 3200 Ci of activity had been dispersed by light variable winds. On 26 September, examination of the geophone records showed six hours of low-level, but fairly continuous, activity before the release. Electrical measurements indicated that most cables were still intact to a depth below the stemming platform. A survey of the ground zero area showed that the seepage came through cracks between the surface conductor and the pad, through cracks in the pad, and through a crack adjacent to the pad around the mousehole (a small hole adjacent to the emplacement hole). To preclude undue radiation exposure or injury from a surprise subsidence, safety measures were instituted. Tritium seepage was suffucient to postpone site activities until a box and pipeline were emplaced to contain and remove the gas. Radiation release modeling and calculations were generally consistent with observations. Plug-hole interaction calculations showed that the alluvium near the bottom of the plug may have been overstressed and that improvements in the design of the plug-medium interface can be made. Experimental studies verified that the surface appearance of the plug core was caused by erosion, but, assuming a normal strength for the plug material, that erosion alone could not account for the disappearance of such a large portion of the stemming platform. Samples from downhole plug experiments show that the plug may have been considerably weaker than had been indicted by quality assurance (QA) samples. 19 references, 32 figures, 10 tables

  6. Allegheny County Toxics Release Inventory

    Data.gov (United States)

    Allegheny County / City of Pittsburgh / Western PA Regional Data Center — The Toxics Release Inventory (TRI) data provides information about toxic substances released into the environment or managed through recycling, energy recovery, and...

  7. Analytical and Clinical Performance Evaluation of the Abbott Architect PIVKA Assay.

    Science.gov (United States)

    Ko, Dae-Hyun; Hyun, Jungwon; Kim, Hyun Soo; Park, Min-Jeong; Kim, Jae-Seok; Park, Ji-Young; Shin, Dong Hoon; Cho, Hyoun Chan

    2018-01-01

    Protein induced by vitamin K absence (PIVKA) is measured using various assays and is used to help diagnose hepatocellular carcinoma. The present study evaluated the analytical and clinical performances of the recently released Abbott Architect PIVKA assay. Precision, linearity, and correlation tests were performed in accordance with the Clinical Laboratory Standardization Institute guidelines. Sample type suitability was assessed using serum and plasma samples from the same patients, and the reference interval was established using sera from 204 healthy individuals. The assay had coefficients of variation of 3.2-3.5% and intra-laboratory variation of 3.6-5.5%. Linearity was confirmed across the entire measurable range. The Architect PIVKA assay was comparable to the Lumipulse PIVKA assay, and the plasma and serum samples provided similar results. The lower reference limit was 13.0 mAU/mL and the upper reference limit was 37.4 mAU/mL. The ability of the Architect PIVKA assay to detect hepatocellular carcinoma was comparable to that of the alpha-fetoprotein test and the Lumipulse PIVKA assay. The Architect PIVKA assay provides excellent analytical and clinical performance, is simple for clinical laboratories to adopt, and has improved sample type suitability that could broaden the assay's utility. © 2018 by the Association of Clinical Scientists, Inc.

  8. Radiosotopic assay and binder therefor

    International Nuclear Information System (INIS)

    Caston, J.D.; Kamen, B.A.

    1976-01-01

    A rapid and less costly radioisotopic assay for measuring the concentration of folate in blood serum is described. This procedure utilizes 3 H-pteroylmonoglutamate, unlabeled 5-methyltetrahydrofolic acid, and a partially purified folate binder, such as for example a folate binder extracted from hog kidney. The procedure involves radioisotopically relating the bound amounts of a labeled folate and a known folate at various concentrations of the known folate in a system containing a predetermined amount of the labeled folate, a predetermined amount of the binder factor for the folates, and a predetermined amount of defolated test serum. 16 claims, 8 drawing figures

  9. Photoresponsive lipid-polymer hybrid nanoparticles for controlled doxorubicin release

    Science.gov (United States)

    Yao, Cuiping; Wu, Ming; Zhang, Cecheng; Lin, Xinyi; Wei, Zuwu; Zheng, Youshi; Zhang, Da; Zhang, Zhenxi; Liu, Xiaolong

    2017-06-01

    Currently, photoresponsive nanomaterials are particularly attractive due to their spatial and temporal controlled drug release abilities. In this work, we report a photoresponsive lipid-polymer hybrid nanoparticle for remote controlled delivery of anticancer drugs. This hybrid nanoparticle comprises three distinct functional components: (i) a poly(D,L-lactide-co-glycolide) (PLGA) core to encapsulate doxorubicin; (ii) a soybean lecithin monolayer at the interface of the core and shell to act as a molecular fence to prevent drug leakage; (iii) a photoresponsive polymeric shell with anti-biofouling properties to enhance nanoparticle stability, which could be detached from the nanoparticle to trigger the drug release via a decrease in the nanoparticle’s stability under light irradiation. In vitro results revealed that this core-shell nanoparticle had excellent light-controlled drug release behavior (76% release with light irradiation versus 10% release without light irradiation). The confocal microscopy and flow cytometry results also further demonstrated the light-controlled drug release behavior inside the cancer cells. Furthermore, a CCK8 assay demonstrated that light irradiation could significantly improve the efficiency of killing cancer cells. Meanwhile, whole-animal fluorescence imaging of a tumor-bearing mouse also confirmed that light irradiation could trigger drug release in vivo. Taken together, our data suggested that a hybrid nanoparticle could be a novel light controlled drug delivery system for cancer therapy.

  10. Antioxidants and the Comet assay.

    Science.gov (United States)

    Cemeli, Eduardo; Baumgartner, Adolf; Anderson, Diana

    2009-01-01

    It is widely accepted that antioxidants, either endogenous or from the diet, play a key role in preserving health. They are able to quench radical species generated in situations of oxidative stress, either triggered by pathologies or xenobiotics, and they protect the integrity of DNA from genotoxicants. Nevertheless, there are still many compounds with unclear or unidentified prooxidant/antioxidant activities. This is of concern since there is an increase in the number of compounds synthesized or extracted from vegetables to which humans might be exposed. Despite the well-established protective effects of fruit and vegetables, the antioxidant(s) responsible have not all been clearly identified. There might also be alternative mechanisms contributing to the protective effects for which a comprehensive description is lacking. In the last two decades, the Comet assay has been extensively used for the investigation of the effects of antioxidants and many reports can be found in the literature. The Comet assay, a relatively fast, simple, and sensitive technique for the analysis of DNA damage in all cell types, has been applied for the screening of chemicals, biomonitoring and intervention studies. In the present review, several of the most well-known antioxidants are considered. These include: catalase, superoxide dismutase, glutathione peroxidase, selenium, iron chelators, melatonin, melanin, vitamins (A, B, C and E), carotenes, flavonoids, isoflavones, tea polyphenols, wine polyphenols and synthetic antioxidants. Investigations showing beneficial as well as non-beneficial properties of the antioxidants selected, either at the in vitro, ex vivo or in vivo level are discussed.

  11. Rotor assembly and assay method

    Science.gov (United States)

    Burtis, C.A.; Johnson, W.F.; Walker, W.A.

    1993-09-07

    A rotor assembly for carrying out an assay includes a rotor body which is rotatable about an axis of rotation, and has a central chamber and first, second, third, fourth, fifth, and sixth chambers which are in communication with and radiate from the central chamber. The rotor assembly further includes a shuttle which is movable through the central chamber and insertable into any of the chambers, the shuttle including a reaction cup carrying an immobilized antigen or an antibody for transport among the chambers. A method for carrying out an assay using the rotor assembly includes moving the reaction cup among the six chambers by passing the cup through the central chamber between centrifugation steps in order to perform the steps of: separating plasma from blood cells, binding plasma antibody or antigen, washing, drying, binding enzyme conjugate, reacting with enzyme substrate and optically comparing the resulting reaction product with unreacted enzyme substrate solution. The movement of the reaction cup can be provided by attaching a magnet to the reaction cup and supplying a moving magnetic field to the rotor. 34 figures.

  12. Cobalt release from inexpensive jewellery

    DEFF Research Database (Denmark)

    Thyssen, Jacob Pontoppidan; Jellesen, Morten Stendahl; Menné, Torkil

    2010-01-01

    . Conclusions: This study showed that only a minority of inexpensive jewellery purchased in Denmark released cobalt when analysed with the cobalt spot test. As fashion trends fluctuate and we found cobalt release from dark appearing jewellery, cobalt release from consumer items should be monitored in the future......Objectives: The aim was to study 354 consumer items using the cobalt spot test. Cobalt release was assessed to obtain a risk estimate of cobalt allergy and dermatitis in consumers who would wear the jewellery. Methods: The cobalt spot test was used to assess cobalt release from all items...

  13. Data transformation methods for multiplexed assays

    Science.gov (United States)

    Tammero, Lance F. Bentley; Dzenitis, John M; Hindson, Benjamin J

    2013-07-23

    Methods to improve the performance of an array assay are described. A correlation between fluorescence intensity-related parameters and negative control values of the assay is determined. The parameters are then adjusted as a function of the correlation. As a result, sensitivity of the assay is improved without changes in its specificity.

  14. Multicentre comparison of a diagnostic assay

    DEFF Research Database (Denmark)

    Waters, Patrick; Reindl, Markus; Saiz, Albert

    2016-01-01

    ) assays in neuromyelitis optica spectrum disorders (NMOSD). METHODS: Coded samples from patients with neuromyelitis optica (NMO) or NMOSD (101) and controls (92) were tested at 15 European diagnostic centres using 21 assays including live (n=3) or fixed cell-based assays (n=10), flow cytometry (n=4...

  15. Capillary Electrophoresis Analysis of Conventional Splicing Assays

    DEFF Research Database (Denmark)

    de Garibay, Gorka Ruiz; Acedo, Alberto; García-Casado, Zaida

    2014-01-01

    of these assays is often challenging. Here, we explore this issue by conducting splicing assays in 31 BRCA2 genetic variants. All variants were assessed by RT-PCR followed by capillary electrophoresis and direct sequencing. If assays did not produce clear-cut outputs (Class-2 or Class-5 according to analytical...

  16. Detection of estrogenic activity in sediment-associated compounds using in vitro reporter gene assays

    NARCIS (Netherlands)

    Legler, J.; Dennekamp, M.; Vethaak, A.D.; Brouwer, A.; Koeman, J.H.; Burg, van der B.; Murk, A.J.

    2002-01-01

    Sediments may be the ultimate sink for persistent (xeno-) estrogenic compounds released into the aquatic environment. Sediment-associated estrogenic potency was measured with an estrogen receptor-mediated luciferase reporter gene (ER-CALUX) assay and compared with a recombinant yeast screen. The

  17. A powerful selection assay for mixture libraries of DNA alkylating agents.

    Science.gov (United States)

    Ham, Young-Wan; Boger, Dale L

    2004-08-04

    A simple and powerful selection assay that permits the separation (rpHPLC), quantitation (ELSD), and identification (ESI-MS) of thermally released adenine adducts derived from duocarmycin analogues is detailed that can establish the most effective DNA alkylating agents in synthetic combinatorial mixtures.

  18. Prevalence of abnormal radiological findings in health care workers with latent tuberculosis infection and correlations with T cell immune response.

    Directory of Open Access Journals (Sweden)

    Rajnish Joshi

    2007-08-01

    Full Text Available More than half of all health care workers (HCWs in high TB-incidence, low and middle income countries are latently infected with tuberculosis (TB. We determined radiological lesions in a cohort of HCWs with latent TB infection (LTBI in India, and determined their association with demographic, occupational and T-cell immune response variables.We obtained chest radiographs of HCWs who had undergone tuberculin skin test (TST and QuantiFERON-TB Gold In Tube (QFT, an interferon-gamma release assay, in a previous cross-sectional study, and were diagnosed to have LTBI because they were positive by either TST or QFT, but had no evidence of clinical disease. Two observers independently interpreted these radiographs using a standardized data form and any discordance between them resolved by a third observer. The radiological diagnostic categories (normal, suggestive of inactive TB, and suggestive of active TB were compared with results of TST, QFT assay, demographic, and occupational covariates.A total of 330 HCWs with positive TST or QFT underwent standard chest radiography. Of these 330, 113 radiographs (34.2% were finally classified as normal, 206 (62.4% had lesions suggestive of inactive TB, and 11 (3.4% had features suggestive of active TB. The mean TST indurations and interferon-gamma levels in the HCWs in these three categories were not significantly different. None of the demographic or occupational covariates was associated with prevalence of inactive TB lesions on chest radiography.In a high TB incidence setting, nearly two-thirds of HCWs with latent TB infection had abnormal radiographic findings, and these findings had no clear correlation with T cell immune responses. Further studies are needed to verify these findings and to identify the causes and prognosis of radiologic abnormalities in health care workers.

  19. Making transuranic assay measurements using modern controllers

    International Nuclear Information System (INIS)

    Kuckertz, T.H.; Caldwell, J.T.; Medvick, P.A.; Kunz, W.E.; Hastings, R.D.

    1987-01-01

    This paper describes methodology and computer-controlled instrumentation developed at the Los Alamos National Laboratory that accurately performs nondestructive assays of large containers bearing transuranic wastes and nonradioactive matrix materials. These assay systems can measure fissile isotopes with 1-mg sensitivity and spontaneous neutron-emitting isotopes at a 10-mg sensitivity. The assays are performed by neutron interrogation, detection, and counting in a custom assay chamber. An International Business Machines Personal Computer (IBM-PC) is used to control the CAMAC-based instrumentation system that acquires the assay data. 6 refs., 7 figs

  20. Non destructive assay (NDA) techniques

    International Nuclear Information System (INIS)

    Mafra Guidicini, Olga; Llacer, Carlos D.; Rojo, Marcelo

    2001-01-01

    In the IAEA Safeguards System the basic verification method used is nuclear material accountancy, with containment and surveillance as important complementary measures. If nuclear material accountancy is to be effective, IAEA inspectors have to make independent measurements to verify declared material quantities. Most of the equipment available to the inspectors is designed to measure gamma rays and/or neutrons emitted by various nuclear materials. Equipment is also available to measure the gross weight of an item containing nuclear material. These types of measurement techniques are generally grouped under the title of nondestructive assay (NDA). The paper describes the NDA techniques and instruments used to verify the total amount of nuclear material held at a nuclear facility. (author)

  1. Assay of cysteine dioxygenase activity

    International Nuclear Information System (INIS)

    Bagley, P.J.; Stipanuk, M.H.

    1990-01-01

    It has been proposed that rat liver contains two cysteine dioxygenase enzymes which convert cysteine to cysteinesulfinic acid, one which is stimulated by NAD + and has a pH optimum of 6.8 and one which is not stimulated by NAD + and has a pH optimum of 9.0. This led the authors to reinvestigate assay conditions for measuring cysteine dioxygenase activity in rat liver homogenate. An HPLC method, using an anion exchange column (Dionex Amino-Pac trademark PA1 (4x250 mm)) was used to separate the [ 35 S]cysteinesulfinic acid produced from [ 35 S]cysteine in the incubation mixture. They demonstrated that inclusion of hydroxylamine prevented further metabolism of cysteinesulfinic acid. which occurred rapidly in the absence of hydroxylamine

  2. Detection of proteins using a colorimetric bio-barcode assay.

    Science.gov (United States)

    Nam, Jwa-Min; Jang, Kyung-Jin; Groves, Jay T

    2007-01-01

    The colorimetric bio-barcode assay is a red-to-blue color change-based protein detection method with ultrahigh sensitivity. This assay is based on both the bio-barcode amplification method that allows for detecting miniscule amount of targets with attomolar sensitivity and gold nanoparticle-based colorimetric DNA detection method that allows for a simple and straightforward detection of biomolecules of interest (here we detect interleukin-2, an important biomarker (cytokine) for many immunodeficiency-related diseases and cancers). The protocol is composed of the following steps: (i) conjugation of target capture molecules and barcode DNA strands onto silica microparticles, (ii) target capture with probes, (iii) separation and release of barcode DNA strands from the separated probes, (iv) detection of released barcode DNA using DNA-modified gold nanoparticle probes and (v) red-to-blue color change analysis with a graphic software. Actual target detection and quantification steps with premade probes take approximately 3 h (whole protocol including probe preparations takes approximately 3 days).

  3. Melamine and Cyanuric Acid do not interfere with Bradford and Ninhydrin assays for protein determination.

    Science.gov (United States)

    Field, Anjalie; Field, Jeffrey

    2010-08-01

    In the fall of 2007 pet food contaminated with melamine and cyanuric acid caused kidney stones in thousands of animals. In the summer of 2008, a more serious outbreak of adulterated dairy food caused the deaths of six infants and sickened about 290,000 children in China. In all cases, melamine was likely added to inflate the apparent protein content of the foods. To determine if we could measure protein without interference from melamine and cyanuric acid we tested these compounds in the Bradford and Ninhydrin assays, two common dye-based assays for protein, as well as by ammonia release, the most common assay used in the food industry. Neither compound was detected in the Ninhydrin and Bradford assays at concentrations of >100 μg/ml. The ammonia assay detected melamine but was inconclusive with respect to cyanuric acid. To develop an accurate test for food that would not detect either chemical as a protein, assays were run on cat food and reconstituted milk powder. The Bradford assay readily measured the protein content of each food, and importantly, the addition of melamine or cyanuric acid to reconstituted milk did not affect the readings. The protein concentrations obtained for reconstituted milk powder were as expected, but those for the cat food were 10 to 30-fold lower, due to its low solubility. We conclude that dye-binding assays can be employed to detect protein in food without interference from melamine and cyanuric acid, thus reducing the incentive to use them as additives.

  4. Q2 - a very low level quantitative and qualitative waste assay and release certification system

    International Nuclear Information System (INIS)

    Bronson, F.L.

    1990-01-01

    Low level radioactive waste disposal is very expensive, especially when all of the handling transportation and documentation costs are included. However for most generators, a large fraction of this low level waste is not contaminated at all, or only slightly so. The paper describes the development and performance of a low level counter that is convenient to use, and that can accurately identify and quantify the radioactivity of any gamma emitter thing that can be placed in a 55 gallon (200 liter) container. These measurement results can be used to verify the absence of radioactivity at a very low levels (10 nCi (370 Bq)/sample), and to identify the nuclides and quantities present, while differentiating against natural radioactivity (Radium, Thorium, Potassium). These results can be used as part of a 10CFR20.302 waste stream exemption program, and thus allow significant savings and a less than 1 year payback at a typical nuclear power plant. The Q1 system is fully shielded to allow it's use in the low level radwaste storage area. The detectors are either 3 Intrinsic Germanium detectors or 2 large NaI detectors. The software is fully automated for simple operation. Correlation factors can be entered to estimate non-gamma emitters from pre-established correlations to other nuclides. Typical Ge detector sensitivities are 8 nCi (300 Bq) LLD for Cs-137 at 0.8 g/cc for a 10 minute count time. NaI detector systems can achieve the same LLD in a 1 minute count. 5 figs., 1 tab

  5. The prevalence of latent Mycobacterium tuberculosis infection based on an interferon-γ release assay

    DEFF Research Database (Denmark)

    Jensen, Andreas V.; Jensen, Lotte; Faurholt-Jepsen, Daniel

    2013-01-01

    Introduction:One third of the world's population is estimated to be latently infected with Mycobacterium tuberculosis (LTBI). Surveys of LTBI are rarely performed in resource poor TB high endemic countries like Tanzania although low-income countries harbor the largest burden of the worlds LTBI....... The primary objective was to estimate the prevalence of LTBI in household contacts of pulmonary TB cases and a group of apparently healthy neighborhood controls in an urban setting of such a country. Secondly we assessed potential impact of LTBI on inflammation by quantitating circulating levels of an acute...

  6. Released radioactivity reducing facility

    International Nuclear Information System (INIS)

    Tanaka, Takeaki.

    1992-01-01

    Upon occurrence of a reactor accident, penetration portions of a reactor container, as a main leakage source from a reactor container, are surrounded by a plurality of gas-tight chambers, the outside of which is surrounded by highly gas-tightly buildings. Branched pipelines of an emergency gas processing system are introduced to each of the gas-tight chambers and they are joined and in communication with an emergency gas processing device. With such a constitution, radioactive materials are prevented from leaking directly from the buildings. Further, pipeline openings of the emergency gas processing facility are disposed in the plurality highly gas-tight penetration chambers. If the radioactive materials are leaked from the reactor to elevate the pressure in the penetration chambers, the radioactive materials are introduced to a filter device in the emergency gas processing facility by way of the branched pipelines, filtered and then released to the atmosphere. Accordingly, the reliability and safety of the system can be improved. (T.M.)

  7. Containment and release management

    International Nuclear Information System (INIS)

    Lehner, J.R.; Pratt, W.T.

    1988-01-01

    Reducing the risk from potentially severe accidents by appropriate accident management strategies is receiving increased attention from the international reactor safety community. Considerable uncertainty still surrounds some of the physical phenomena likely to occur during a severe accident. The USNRC, in developing its research plan for accident management, wants to ensure that both the developers and implementers of accident management strategies are aware of the uncertainty associated with the plant operators' ability to correctly diagnose an accident, as well as the uncertainties associated with various preventive and mitigative strategies. The use of a particular accident management strategy can have both positive and negative effects on the status of a plant and these effects must be carefully weighed before a particular course of action is chosen and implemented. By using examples of severe accident scenarios, initial insights are presented here regarding the indications plant operators may have to alert them to particular accident states. Insights are also offered on the various management actions operators and plant technical staff might pursue for particular accident situations and the pros and cons associated with such actions. The examples given are taken for the most part from the containment and release phase of accident management, since this is the current focus of the effort in the accident management area at Brookhaven National Laboratory. 2 refs

  8. Released radioactivity reducing device

    International Nuclear Information System (INIS)

    Miyamoto, Yumi.

    1995-01-01

    A water scrubber is disposed in a scrubber tank and a stainless steel fiber filter is disposed above the water scrubber. The upper end of the scrubber tank is connected by way of a second bent tube to a capturing vessel incorporating a moisture removing layer and an activated carbon filter. The exit of the capturing vessel is connected to a stack. Upon occurrence of an accident of a BWR-type power plant, gases containing radioactive materials released from a reactor container are discharged into the water scrubber from a first bent tube through a venturi tube nozzle, and water soluble and aerosol-like radioactive materials are captured in the water. Aerosol and splashes of water droplets which can not be captured thoroughly by the water scrubber are captured by the stainless steel fiber filter. Gases passing through the scrubber tank are introduced to a capturing vessel through a second bent tube, and organic iodine is captured by the activated carbon filter. (I.N.)

  9. COMMERCIAL SNF ACCIDENT RELEASE FRACTIONS

    Energy Technology Data Exchange (ETDEWEB)

    S.O. Bader

    1999-10-18

    The purpose of this design analysis is to specify and document the total and respirable fractions for radioactive materials that are released from an accident event at the Monitored Geologic Repository (MGR) involving commercial spent nuclear fuel (CSNF) in a dry environment. The total and respirable release fractions will be used to support the preclosure licensing basis for the MGR. The total release fraction is defined as the fraction of total CSNF assembly inventory, typically expressed as an activity inventory (e.g., curies), of a given radionuclide that is released to the environment from a waste form. The radionuclides are released from the inside of breached fuel rods (or pins) and from the detachment of radioactive material (crud) from the outside surfaces of fuel rods and other components of fuel assemblies. The total release fraction accounts for several mechanisms that tend to retain, retard, or diminish the amount of radionuclides that are available for transport to dose receptors or otherwise can be shown to reduce exposure of receptors to radiological releases. The total release fraction includes a fraction of airborne material that is respirable and could result in inhalation doses. This subset of the total release fraction is referred to as the respirable release fraction. Potential accidents may involve waste forms that are characterized as either bare (unconfined) fuel assemblies or confined fuel assemblies. The confined CSNF assemblies at the MGR are contained in shipping casks, canisters, or disposal containers (waste packages). In contrast to the bare fuel assemblies, the container that confines the fuel assemblies has the potential of providing an additional barrier for diminishing the total release fraction should the fuel rod cladding breach during an accident. However, this analysis will not take credit for this additional bamer and will establish only the total release fractions for bare unconfined CSNF assemblies, which may however be

  10. COMMERCIAL SNF ACCIDENT RELEASE FRACTIONS

    International Nuclear Information System (INIS)

    S.O. Bader

    1999-01-01

    The purpose of this design analysis is to specify and document the total and respirable fractions for radioactive materials that are released from an accident event at the Monitored Geologic Repository (MGR) involving commercial spent nuclear fuel (CSNF) in a dry environment. The total and respirable release fractions will be used to support the preclosure licensing basis for the MGR. The total release fraction is defined as the fraction of total CSNF assembly inventory, typically expressed as an activity inventory (e.g., curies), of a given radionuclide that is released to the environment from a waste form. The radionuclides are released from the inside of breached fuel rods (or pins) and from the detachment of radioactive material (crud) from the outside surfaces of fuel rods and other components of fuel assemblies. The total release fraction accounts for several mechanisms that tend to retain, retard, or diminish the amount of radionuclides that are available for transport to dose receptors or otherwise can be shown to reduce exposure of receptors to radiological releases. The total release fraction includes a fraction of airborne material that is respirable and could result in inhalation doses. This subset of the total release fraction is referred to as the respirable release fraction. Potential accidents may involve waste forms that are characterized as either bare (unconfined) fuel assemblies or confined fuel assemblies. The confined CSNF assemblies at the MGR are contained in shipping casks, canisters, or disposal containers (waste packages). In contrast to the bare fuel assemblies, the container that confines the fuel assemblies has the potential of providing an additional barrier for diminishing the total release fraction should the fuel rod cladding breach during an accident. However, this analysis will not take credit for this additional bamer and will establish only the total release fractions for bare unconfined CSNF assemblies, which may however be

  11. The assay of glucocerebrosidase activity using the natural substrate

    International Nuclear Information System (INIS)

    Strasberg, P.M.; Lowden, J.A.

    1982-01-01

    The authors have examined eight published methods for the assay of glucocerebrosidase using the natural substrate glucocerebroside and tabulated the variable components and conditions while comparing these methods to their own. In each case assays were performed using a 8000 fold, partially purified human placental enzyme, having a pH optimum of 5.4-5.6 and a Ksub(m) of approximately 0.60 mmol per liter. Results were obtained by following the release of radioactive ceramide or of unlabelled glucose. In many cases published results had been based on a one- or two-hour incubation time. Apparent specific activities varied over a 70-fold difference between the various procedures. When the authors measured activities from the linear (15-30 min) portion of rate curves the values increased by 1.4 to 3 times but still ranged from 6x10 3 -180x10 3 nmol. mg -1 protein h -1 . They obtained maximum velocity using 1.2 mmol glucocerebroside, 0.5% (w/v) crude taurocholate and 2 μg enzyme protein/ml. Specific activities reported from different laboratories are not directly comparable. Conditions for assay should be optimized for the enzyme preparation to be studied. (Auth.)

  12. Comparative assessment of in vitro release kinetics of calcitonin polypeptide from biodegradable microspheres.

    Science.gov (United States)

    Prabhu, Sunil; Sullivan, Jennifer L; Betageri, Guru V

    2002-01-01

    The objective of our study was to compare the in vitro release kinetics of a sustained-release injectable microsphere formulation of the polypeptide drug, calcitonin (CT), to optimize the characteristics of drug release from poly-(lactide-co-glycolide) (PLGA) copolymer biodegradable microspheres. A modified solvent evaporation and double emulsion technique was used to prepare the microspheres. Release kinetic studies were carried out in silanized tubes and dialysis bags, whereby microspheres were suspended and incubated in phosphate buffered saline, sampled at fixed intervals, and analyzed for drug content using a modified Lowry protein assay procedure. An initial burst was observed whereby about 50% of the total dose of the drug was released from the microspheres within 24 hr and 75% within 3 days. This was followed by a period of slow release over a period of 3 weeks in which another 10-15% of drug was released. Drug release from the dialysis bags was more gradual, and 50% CT was released only after 4 days and 75% after 12 days of release. Scanning electron micrographs revealed spherical particles with channel-like structures and a porous surface after being suspended in an aqueous solution for 5 days. Differential scanning calorimetric studies revealed that CT was present as a mix of amorphous and crystalline forms within the microspheres. Overall, these studies demonstrated that sustained release of CT from PLGA microspheres over a 3-week period is feasible and that release of drug from dialysis bags was more predictable than from tubes.

  13. Underground water stress release models

    Science.gov (United States)

    Li, Yong; Dang, Shenjun; Lü, Shaochuan

    2011-08-01

    The accumulation of tectonic stress may cause earthquakes at some epochs. However, in most cases, it leads to crustal deformations. Underground water level is a sensitive indication of the crustal deformations. We incorporate the information of the underground water level into the stress release models (SRM), and obtain the underground water stress release model (USRM). We apply USRM to the earthquakes occurred at Tangshan region. The analysis shows that the underground water stress release model outperforms both Poisson model and stress release model. Monte Carlo simulation shows that the simulated seismicity by USRM is very close to the real seismicity.

  14. Expert system for transuranic waste assay

    Energy Technology Data Exchange (ETDEWEB)

    Zoolalian, M.L.; Gibbs, A.; Kuhns, J.D.

    1989-01-01

    Transuranic wastes are generated at the Savannah River Site (SRS) as a result of routine production of nuclear materials. These wastes contain Pu-238 and Pu-239 and are placed into lined 55-gallon waste drums. The drums are placed on monitored storage pads pending shipment to the Waste Isolation Pilot Plant in New Mexico. A passive-active neutron (PAN) assay system is used to determine the mass of the radioactive material within the waste drums. Assay results are used to classify the wastes as either low-level or transuranic (TRU). During assays, the PAN assay system communicates with an IBM-AT computer. A Fortran computer program, called NEUT, controls and performs all data analyses. Unassisted, the NEUT program cannot adequately interpret assay results. To eliminate this limitation, an expert system shell was used to write a new algorithm, called the Transuranic Expert System (TRUX), to drive the NEUT program and add decision making capabilities for analysis of the assay results. The TRUX knowledge base was formulated by consulting with human experts in the field of neutron assay, by direct experimentation on the PAN assay system, and by observing operations on a daily basis. TRUX, with its improved ability to interpret assay results, has eliminated the need for close supervision by a human expert, allowing skilled technicians to operate the PAN assay system. 4 refs., 1 fig., 4 tabs.

  15. Expert system for transuranic waste assay

    International Nuclear Information System (INIS)

    Zoolalian, M.L.; Gibbs, A.; Kuhns, J.D.

    1989-01-01

    Transuranic wastes are generated at the Savannah River Site (SRS) as a result of routine production of nuclear materials. These wastes contain Pu-238 and Pu-239 and are placed into lined 55-gallon waste drums. The drums are placed on monitored storage pads pending shipment to the Waste Isolation Pilot Plant in New Mexico. A passive-active neutron (PAN) assay system is used to determine the mass of the radioactive material within the waste drums. Assay results are used to classify the wastes as either low-level or transuranic (TRU). During assays, the PAN assay system communicates with an IBM-AT computer. A Fortran computer program, called NEUT, controls and performs all data analyses. Unassisted, the NEUT program cannot adequately interpret assay results. To eliminate this limitation, an expert system shell was used to write a new algorithm, called the Transuranic Expert System (TRUX), to drive the NEUT program and add decision making capabilities for analysis of the assay results. The TRUX knowledge base was formulated by consulting with human experts in the field of neutron assay, by direct experimentation on the PAN assay system, and by observing operations on a daily basis. TRUX, with its improved ability to interpret assay results, has eliminated the need for close supervision by a human expert, allowing skilled technicians to operate the PAN assay system. 4 refs., 1 fig., 4 tabs

  16. Radiometric assays for glycerol, glucose, and glycogen

    International Nuclear Information System (INIS)

    Bradley, D.C.; Kaslow, H.R.

    1989-01-01

    We have developed radiometric assays for small quantities of glycerol, glucose and glycogen, based on a technique described by Thorner and Paulus for the measurement of glycerokinase activity. In the glycerol assay, glycerol is phosphorylated with [32P]ATP and glycerokinase, residual [32P]ATP is hydrolyzed by heating in acid, and free [32P]phosphate is removed by precipitation with ammonium molybdate and triethylamine. Standard dose-response curves were linear from 50 to 3000 pmol glycerol with less than 3% SD in triplicate measurements. Of the substances tested for interference, only dihydroxyacetone gave a slight false positive signal at high concentration. When used to measure glycerol concentrations in serum and in media from incubated adipose tissue, the radiometric glycerol assay correlated well with a commonly used spectrophotometric assay. The radiometric glucose assay is similar to the glycerol assay, except that glucokinase is used instead of glycerokinase. Dose response was linear from 5 to 3000 pmol glucose with less than 3% SD in triplicate measurements. Glucosamine and N-acetylglucosamine gave false positive signals when equimolar to glucose. When glucose concentrations in serum were measured, the radiometric glucose assay agreed well with hexokinase/glucose-6-phosphate dehydrogenase (H/GDH)-based and glucose oxidase/H2O2-based glucose assays. The radiometric method for glycogen measurement incorporates previously described isolation and digestion techniques, followed by the radiometric assay of free glucose. When used to measure glycogen in mouse epididymal fat pads, the radiometric glycogen assay correlated well with the H/GDH-based glycogen assay. All three radiometric assays offer several practical advantages over spectral assays

  17. Harmonization of radiobiological assays: why and how?

    International Nuclear Information System (INIS)

    Prasanna, Pataje G.

    2014-01-01

    The International Atomic Energy Agency has made available a technical manual for cytogenetic biodosimetry assays (dicentric chromosome aberration (DCA) and cytokinesis-block micronucleus (CBMN) assays) used for radiation dose assessment in radiation accidents. The International Standardization Organization, which develops standards and guidelines, also provides an avenue for laboratory accreditation, has developed guidelines and recommendations for performing cytogenetic biodosimetry assays. Harmonization of DCA and CBMN assays, has improved their accuracy. Double-blinded inter-laboratory comparison studies involving several networks have further validated DCA and CBMN assays and improved the confidence in their potential use for radiation dose assessment in mass casualties. This kind of international harmonization is lacking for pre-clinical radiobiology assays. The widely used pre-clinical assays that are relatively important to set stage for clinical trials include clonogenic assays, flow-cytometry assays, apoptotic assays, and tumor regression and growth delay assays. However, significant inter-laboratory variations occur with respect to data among laboratories. This raises concerns on the reliability and reproducibility of preclinical data that drives further development and translation. Lack of reproducibility may stem from a variety of factors such as poor scientist training, less than optimal experimental design, inadequate description of methodology, and impulse to publish only the positive data etc. Availability of technical manuals, standard operating procedures, accreditation avenues for laboratories performing such assays, inter-laboratory comparisons, and use of standardized protocols are necessary to enhance reliability and reproducibility. Thus, it is important that radiobiological assays are harmonized for laboratory protocols to ensure successful translation of pre-clinical research on radiation effect modulators to help design clinic trials with

  18. Flash release an alternative for releasing complex MEMS devices

    NARCIS (Netherlands)

    Deladi, S.; Krijnen, Gijsbertus J.M.; Elwenspoek, Michael Curt

    2004-01-01

    A novel time-saving and cost-effective release technique has been developed and is described. The physical nature of the process is explained in combination with experimental observations. The results of the flash release process are compared with those of freeze-drying and supercritical CO2

  19. The self-antigen, thyroglobulin, induces antigen-experienced CD4+ T cells from healthy donors to proliferate and promote production of the regulatory cytokine, interleukin-10, by monocytes

    DEFF Research Database (Denmark)

    Nielsen, Claus H; Galdiers, Marcel P; Hedegaard, Chris J

    2010-01-01

    . Whereas TT induced pro-inflammatory cytokines [interleukin-2 (IL-2)/interferon-gamma (IFN-gamma)/IL-4/IL-5], TG evoked persistent release of the regulatory IL-10. Some donors, however, also responded with late IFN-gamma production, suggesting that the regulation by IL-10 could be overridden. Although...

  20. Automated amperometric plutonium assay system

    International Nuclear Information System (INIS)

    Burt, M.C.

    1985-01-01

    The amperometric titration for plutonium assay has been used in the nuclear industry for over twenty years and has been in routine use at the Hanford Engineering Development Laboratory since 1976 for the analysis of plutonium oxide and mixed oxide fuel material for the Fast Flux Test Facility. It has proven itself to be an accurate and reliable method. The method may be used as a direct end point titration or an excess of titrant may be added and a back titration performed to aid in determination of the end point. Due to the slowness of the PuVI-FeII reaction it is difficult to recognize when the end point is being approached and is very time consuming if the current is allowed to decay to the residual value after each titrant addition. For this reason the back titration in which the rapid FeII-CrVI reaction occurs is used by most laboratories. The back titration is performed by the addition of excess ferrous solution followed by two measured aliquots of standard dichromate with measurement of cell current after each addition

  1. TRU assay system and measurements

    International Nuclear Information System (INIS)

    Brodzinski, R.L.

    1984-02-01

    The measurement of the transuranic content of nuclear products or process residues has become increasingly important for the recovery of fissionable material from spent fuel elements, the identification of commercial fuel elements which have not yet reached full burnup, the measurement and recovery of transuranics from discarded or stored waste materials, the determination of the transuranic content in high gamma activity waste material scheduled for disposal, compliance with 10CFR61 by land burial operators/shippers, and the satisfaction of accountability requirements. Active neutron interrogation techniques measure either the prompt neutrons or the beta delayed neutrons from fission products following induced fission. These techniques normally only measure fissile transuranics ( 235 U, 239 Pu, and 241 Pu) and are commonly applied only to contact handleable waste. Passive neutron interrogation techniques, on the other hand, are capable of measuring all transuranics except 235 U with adequate sensitivity and will work on both contact handleable and high gamma activity wastes. Since the passive techniques are senstitive to a wider spectrum of transuranic isotopes than the active techniques, substantially less complex and less expensive than the active systems, and they have proven techniques for measuring small quantities of TRU in high gamma activity packages, the passive neutron TRU assay technology was chosen for development into the instruments discussed in this paper

  2. Micronucleus assay for radiation workers

    International Nuclear Information System (INIS)

    Balasem, A.N.; Ali, A.S.K.

    1997-01-01

    Micronucleus assay was performed on 49 radiation workers and 22 healthy volunteers. Radiation workers were subdivided into two groups according to their employments durations in the radiation field. Group a consisted of 18 radiation workers who have been in this work between 5 and 22 years. Group b included 31 employees who have been classified as radiation workers for 1 to 4.5 years. Statistical analysis showed significant variations between the yields of micronuclei in groups A and B as well as between group A and a group of healthy controls. Meanwhile no significant difference was noticed between the yields of micronuclei in group B and the corresponding values in the healthy controls. The possible effect of age in the induction of micronuclei was discussed and a comparison with the yield of chromosomal aberrations was described. It seems that cytokinesis- blocking method may be used to detect the radiation-induced micronuclei in workers exposed occupationally to ionizing radiation in levels below the maximum permissible limit of 0.05 Sv per year

  3. PAME: plasmonic assay modeling environment

    Directory of Open Access Journals (Sweden)

    Adam Hughes

    2015-08-01

    Full Text Available Plasmonic assays are an important class of optical sensors that measure biomolecular interactions in real-time without the need for labeling agents, making them especially well-suited for clinical applications. Through the incorporation of nanoparticles and fiberoptics, these sensing systems have been successfully miniaturized and show great promise for in-situ probing and implantable devices, yet it remains challenging to derive meaningful, quantitative information from plasmonic responses. This is in part due to a lack of dedicated modeling tools, and therefore we introduce PAME, an open-source Python application for modeling plasmonic systems of bulk and nanoparticle-embedded metallic films. PAME combines aspects of thin-film solvers, nanomaterials and fiber-optics into an intuitive graphical interface. Some of PAME’s features include a simulation mode, a database of hundreds of materials, and an object-oriented framework for designing complex nanomaterials, such as a gold nanoparticles encased in a protein shell. An overview of PAME’s theory and design is presented, followed by example simulations of a fiberoptic refractometer, as well as protein binding to a multiplexed sensor composed of a mixed layer of gold and silver colloids. These results provide new insights into observed responses in reflectance biosensors.

  4. High-throughput screening assay of hepatitis C virus helicase inhibitors using fluorescence-quenching phenomenon

    International Nuclear Information System (INIS)

    Tani, Hidenori; Akimitsu, Nobuyoshi; Fujita, Osamu; Matsuda, Yasuyoshi; Miyata, Ryo; Tsuneda, Satoshi; Igarashi, Masayuki; Sekiguchi, Yuji; Noda, Naohiro

    2009-01-01

    We have developed a novel high-throughput screening assay of hepatitis C virus (HCV) nonstructural protein 3 (NS3) helicase inhibitors using the fluorescence-quenching phenomenon via photoinduced electron transfer between fluorescent dyes and guanine bases. We prepared double-stranded DNA (dsDNA) with a 5'-fluorescent-dye (BODIPY FL)-labeled strand hybridized with a complementary strand, the 3'-end of which has guanine bases. When dsDNA is unwound by helicase, the dye emits fluorescence owing to its release from the guanine bases. Our results demonstrate that this assay is suitable for quantitative assay of HCV NS3 helicase activity and useful for high-throughput screening for inhibitors. Furthermore, we applied this assay to the screening for NS3 helicase inhibitors from cell extracts of microorganisms, and found several cell extracts containing potential inhibitors.

  5. Assay development status report for total cyanide

    International Nuclear Information System (INIS)

    Simpson, B.C.; Jones, T.E.; Pool, K.H.

    1993-02-01

    A validated cyanide assay that is applicable to a variety of tank waste matrices is necessary to resolve certain waste tank safety issues and for purposes of overall waste characterization. The target for this effort is an assay with an applicable range of greater than 1,000 ppM (0.10 wt%) total cyanide and a confidence level greater than 80%. Figure 1 illustrates the operating regime of the proposed cyanide assay method. The Assay Development Status Report for Total Cyanide will summarize the past experience with cyanide analyses on-tank waste matrices and will rate the status of the analytical methods used to assay total cyanide (CN - ion) in the tank waste matrices as acceptable or unacceptable. This paper will also briefly describe the current efforts for improving analytical resolution of the assays and the attempts at speciation

  6. Linearization of the Bradford Protein Assay

    OpenAIRE

    Ernst, Orna; Zor, Tsaffrir

    2010-01-01

    Determination of microgram quantities of protein in the Bradford Coomassie brilliant blue assay is accomplished by measurement of absorbance at 590 nm. This most common assay enables rapid and simple protein quantification in cell lysates, cellular fractions, or recombinant protein samples, for the purpose of normalization of biochemical measurements. However, an intrinsic nonlinearity compromises the sensitivity and accuracy of this method. It is shown that under standard assay conditions, t...

  7. New automated pellet/powder assay system

    International Nuclear Information System (INIS)

    Olsen, R.N.

    1975-01-01

    This paper discusses an automated, high precision, pellet/ powder assay system. The system is an active assay system using a small isotopic neutron source and a coincidence detection system. The handling of the pellet powder samples has been automated and a programmable calculator has been integrated into the system to provide control and data analysis. The versatile system can assay uranium or plutonium in either active or passive modes

  8. Matrix effects of TRU [transuranic] assays using the SWEPP PAN assay system

    International Nuclear Information System (INIS)

    Smith, J.R.

    1990-08-01

    The Drum Assay System (DAS) at the Stored Waste Experimental Pilot Plant (SWEPP) is a second-generation active-passive neutron assay system. It has been used to assay over 5000 208-liter drums of transuranic waste from the Rocky Flats Plant (RFP). Data from these assays have been examined and compared with the assays performed at Rocky Flats, mainly utilize counting of 239 Pu gamma rays. For the most part the passive assays are in very good agreement with the Rocky Flats assays. The active assays are strongly correlated with the results of the other two methods, but require matrix-dependent correction factors beyond those provided by the system itself. A set of matrix-dependent correction factors has been developed from the study of the assay results. 3 refs., 4 figs., 3 tabs

  9. Sustained release of radioprotective agents

    International Nuclear Information System (INIS)

    Shani, J.

    1980-11-01

    New pharmaceutical formulations for the sustained release into the G.I. tract of radioprotective agents have been developed by the authors. The experimental method initially consisted in the production of methylcellulose microcapsules. This method failed apparently because of the premature ''explosion'' of the microcapsules and the consequent premature release of massive amounts of the drug. A new method has been developed which consists in drying and pulverising cysteamine and cysteine preparations, mixing them in various proportions with stearic acid and ethylcellulose as carriers. The mixture is then compressed into cylindrical tablets at several pressure values and the leaching rate of the radioprotective agents is then measured by spectrophotometry. The relation between the concentration of the active drug and its rate of release, and the effect on the release rate of the pressure applied to the tablet during its formation were also investigated. Results indicating that the release rate was linearly related to the square root of ''t'' seem to be in agreement with what is predictable, according to Higuchi's equation, save for the very initial and terminal phases. A clear correlation was also established between the stearic acid/ethylcellulose ratios and the release of 20% cysteine, namely a marked decrease in the rate of cysteine release was observed with increasing concentrations of stearic acid. Finally, it was observed that a higher formation pressure results in quicker release of the drug

  10. Press Oil Final Release Survey

    Energy Technology Data Exchange (ETDEWEB)

    Whicker, Jeffrey Jay [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Ruedig, Elizabeth [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2017-05-11

    There are forty-eight 55 gallon barrels filled with hydraulic oil that are candidates for release and recycle. This oil needs to be characterized prior to release. Principles of sampling as provided in MARSAME/MARSSIM approaches were used as guidance for sampling.

  11. Workload Control with Continuous Release

    NARCIS (Netherlands)

    Phan, B. S. Nguyen; Land, M. J.; Gaalman, G. J. C.

    2009-01-01

    Workload Control (WLC) is a production planning and control concept which is suitable for the needs of make-to-order job shops. Release decisions based on the workload norms form the core of the concept. This paper develops continuous time WLC release variants and investigates their due date

  12. A deadenylase assay by size-exclusion chromatography.

    Science.gov (United States)

    He, Guang-Jun; Yan, Yong-Bin

    2012-01-01

    The shortening of the 3'-end poly(A) tail, also called deadenylation, is crucial to the regulation of mRNA processing, transportation, translation and degradation. The deadenylation process is achieved by deadenylases, which specifically catalyze the removal of the poly(A) tail at the 3'-end of eukaryotic mRNAs and release 5'-AMP as the product. To achieve their physiological functions, all deadenylases have numerous binding partners that may regulate their catalytic properties or recruit them into various protein complexes. To study the effects of various partners, it is important to develop new deadenylase assay that can be applied either in vivo or in vitro. In this research, we developed the deadenylase assay by the size-exclusion chromatography (SEC) method. The SEC analysis indicated that the poly(A) or oligo(A) substrate and the product AMP could be successfully separated and quantified. The enzymatic parameters of deadenylase could be obtained by quantifying the AMP generation. When using the commercial poly(A) as the substrate, a biphasic catalytic process was observed, which might correlate to the two distinct states of poly(A) in the commercial samples. Different lots of commercial poly(A) had dissimilar size distributions and were dissimilar in response to the degradation of deadenylase. The deadenylation pattern, processive or distributive, could also be investigated using the SEC assay by monitoring the status of the substrate and the generation kinetics of AMP and A2. The SEC assay was applicable to both simple samples using the purified enzyme and complex enzyme reaction conditions such as using protein mixtures or crude cell extracts as samples. The influence of solutes with absorption at 254 nm could be successfully eliminated by constructing the different SEC profiles.

  13. Multicenter Evaluation of Cystatin C Measurement after Assay Standardization.

    Science.gov (United States)

    Bargnoux, Anne-Sophie; Piéroni, Laurence; Cristol, Jean-Paul; Kuster, Nils; Delanaye, Pierre; Carlier, Marie-Christine; Fellahi, Soraya; Boutten, Anne; Lombard, Christine; González-Antuña, Ana; Delatour, Vincent; Cavalier, Etienne

    2017-04-01

    Since 2010, a certified reference material ERM-DA471/IFCC has been available for cystatin C (CysC). This study aimed to assess the sources of uncertainty in results for clinical samples measured using standardized assays. This evaluation was performed in 2015 and involved 7 clinical laboratories located in France and Belgium. CysC was measured in a panel of 4 serum pools using 8 automated assays and a candidate isotope dilution mass spectrometry reference measurement procedure. Sources of uncertainty (imprecision and bias) were evaluated to calculate the relative expanded combined uncertainty for each CysC assay. Uncertainty was judged against the performance specifications derived from the biological variation model. Only Siemens reagents on the Siemens systems and, to a lesser extent, DiaSys reagents on the Cobas system, provided results that met the minimum performance criterion calculated according to the intraindividual and interindividual biological variations. Although the imprecision was acceptable for almost all assays, an increase in the bias with concentration was observed for Gentian reagents, and unacceptably high biases were observed for Abbott and Roche reagents on their own systems. This comprehensive picture of the market situation since the release of ERM-DA471/IFCC shows that bias remains the major component of the combined uncertainty because of possible problems associated with the implementation of traceability. Although some manufacturers have clearly improved their calibration protocols relative to ERM-DA471, most of them failed to meet the criteria for acceptable CysC measurements. © 2016 American Association for Clinical Chemistry.

  14. 233U Assay A Neutron NDA System

    Energy Technology Data Exchange (ETDEWEB)

    Hensley, D.C.; Lucero, A.J.; Pierce, L.

    1998-11-17

    The assay of highly enriched {sup 233}U material presents some unique challenges. Techniques which apply to the assay of materials of Pu or enriched {sup 235}U do not convert easily over to the assay of {sup 233}U. A specialized neutron assay device is being fabricated to exploit the singles neutron signal, the weak correlated neutron signal, and an active correlated signal. These pieces of information when combined with {gamma} ray isotopics information should give a good overall determination of {sup 233}U material now stored in bldg. 3019 at the Oak Ridge National Laboratory.

  15. 233U Assay A Neutron NDA System

    International Nuclear Information System (INIS)

    Hensley, D.C.; Lucero, A.J.; Pierce, L.

    1998-01-01

    The assay of highly enriched 233 U material presents some unique challenges. Techniques which apply to the assay of materials of Pu or enriched 235 U do not convert easily over to the assay of 233 U. A specialized neutron assay device is being fabricated to exploit the singles neutron signal, the weak correlated neutron signal, and an active correlated signal. These pieces of information when combined with γ ray isotopics information should give a good overall determination of 233 U material now stored in bldg. 3019 at the Oak Ridge National Laboratory

  16. Safeguards and Non-destructive Assay

    International Nuclear Information System (INIS)

    Carchon, R.; Bruggeman, M.

    2001-01-01

    SCK-CEN's programme on safeguards and non-destructive assay includes: (1) various activities to assure nuclear materials accountancy; (2) contributes to the implementation of Integrated Safeguards measures in Belgium and to assist the IAEA through the Belgian Support Programme; (3) renders services to internal and external customers in the field of safeguards; (4) improves passive neutron coincidence counting techniques for waste assay and safeguards verification measurements by R and D on correlation algorithms implemented via software or dedicated hardware; (5) improves gamma assay techniques for waste assay by implementing advanced scanning techniques and different correlation algorithms; and (6) develops numerical calibration techniques. Major achievements in these areas in 2000 are reported

  17. Assay-specific decision limits for two new automated parathyroid hormone and 25-hydroxyvitamin D assays.

    Science.gov (United States)

    Souberbielle, Jean-Claude; Fayol, Véronique; Sault, Corinne; Lawson-Body, Ethel; Kahan, André; Cormier, Catherine

    2005-02-01

    The recent development of nonradioactive automated assays for serum parathyroid hormone (PTH) and 25-hydroxyvitamin D (25OHD) has made measurement of these two hormones possible in many laboratories. In this study, we compared two new assays for PTH and 25OHD adapted on an automated analyzer, the LIAISON, with two manual immunoassays used worldwide. We studied 228 osteoporotic patients, 927 healthy individuals, 38 patients with primary hyperparathyroidism, and 167 hemodialyzed patients. Serum PTH was measured with the Allegro and the LIAISON assays, and 25OHD was measured with DiaSorin RIA and the LIAISON assay. Regression analysis was used to calculate decision thresholds for the LIAISON assays that were equivalent to those of the Allegro PTH and DiaSorin 25OHD assays. The 25OHD concentrations obtained with the LIAISON assay and the RIA in osteoporotic patients were well correlated (r = 0.83; P 50 nmol/L as eligible for the reference population for the LIAISON PTH assay. In this group, the 3rd-97th percentile interval for LIAISON PTH was 3-51 ng/L. Considering upper reference limits of 46 and 51 ng/L for the Allegro and LIAISON assays, respectively, the frequency of above-normal PTH concentrations in patients with primary hyperparathyroidism was similar in both assays. Regression analysis between serum PTH measured by the Allegro and LIAISON assays in 167 hemodialyzed patients and the corresponding Bland-Altman analysis of these data suggest that the LIAISON PTH assay tends to read higher than the Allegro assay at low concentrations but lower at high concentrations (>300 ng/L). Because clinical decision limits for both PTH and 25OHD should be assay specific, we propose equivalences between these assays and two manual assays used worldwide. These assay-specific decision limits should help potential users of the LIAISON PTH and 25OHD assays.

  18. Toxic releases from power plants

    International Nuclear Information System (INIS)

    Rubin, E.S.

    1999-01-01

    Beginning in 1998, electric power plants burning coal or oil must estimate and report their annual releases of toxic chemicals listed in the Toxics Release Inventory (TRI) published by the US Environmental Protection Agency (EPA). This paper identifies the toxic chemicals of greatest significance for the electric utility sector and develops quantitative estimates of the toxic releases reportable to the TRI for a representative coal-fired power plant. Key factors affecting the magnitude and types of toxic releases for individual power plants also are discussed. A national projection suggests that the magnitude of electric utility industry releases will surpass those of the manufacturing industries which current report to the TRI. Risk communication activities at the community level will be essential to interpret and provide context for the new TRI results

  19. Enzyme activity assay of glycoprotein enzymes based on a boronate affinity molecularly imprinted 96-well microplate.

    Science.gov (United States)

    Bi, Xiaodong; Liu, Zhen

    2014-12-16

    Enzyme activity assay is an important method in clinical diagnostics. However, conventional enzyme activity assay suffers from apparent interference from the sample matrix. Herein, we present a new format of enzyme activity assay that can effectively eliminate the effects of the sample matrix. The key is a 96-well microplate modified with molecularly imprinted polymer (MIP) prepared according to a newly proposed method called boronate affinity-based oriented surface imprinting. Alkaline phosphatase (ALP), a glycoprotein enzyme that has been routinely used as an indicator for several diseases in clinical tests, was taken as a representative target enzyme. The prepared MIP exhibited strong affinity toward the template enzyme (with a dissociation constant of 10(-10) M) as well as superb tolerance for interference. Thus, the enzyme molecules in a complicated sample matrix could be specifically captured and cleaned up for enzyme activity assay, which eliminated the interference from the sample matrix. On the other hand, because the boronate affinity MIP could well retain the enzymatic activity of glycoprotein enzymes, the enzyme captured by the MIP was directly used for activity assay. Thus, additional assay time and possible enzyme or activity loss due to an enzyme release step required by other methods were avoided. Assay of ALP in human serum was successfully demonstrated, suggesting a promising prospect of the proposed method in real-world applications.

  20. Evaluation of assays for quantification of DNA in canine plasma as an indirect marker of NETosis.

    Science.gov (United States)

    Smith, Stephanie A; Lawson, Corinne M; McMichael, Maureen A; Jung, Katrina; O'Brien, Mauria; Achiel, Ron

    2017-06-01

    Neutrophil extracellular traps (NET), consisting of a filamentous DNA/chromatin-histone scaffold originating from neutrophils are part of the innate immune response, may be released under a variety of inflammatory conditions and are associated with an increased risk for thrombosis. The purpose of this study was to evaluate a SYTOX green fluorescence assay and a histone-DNA complex (hisDNA) ELISA for quantification of NET-related DNA in canine plasma. The influence of variations in blood sample handling on assay results was tested. Accuracy of the SYTOX green fluorescence and the hisDNA ELISA was evaluated with dilutional linearity using serial dilutions. Interference was assessed by addition of purified bilirubin or hemoglobin. Precision was determined by calculating the intra- and inter-assay CV. Preanalytic sample handling did not influence DNA measurements by either assay. Citrate and EDTA plasma samples were equivalent. For the DNA fluorescence assay, dilutional linearity was poor due to autofluorescence, which was corrected by addition of canine plasma to the diluent. The presence of bilirubin and hemoglobin also increased autofluorescence, and resulted in falsely low concentrations of DNA. On the hisDNA ELISA, pigmentemia had no effect. Both assays as modified in this study are suitable for measuring DNA in canine EDTA or citrate plasma. However, performance of the fluorescence assay was impacted by pigmentemia, and it was less sensitive than the ELISA in detecting the presence of nucleosome material in the plasma. © 2017 American Society for Veterinary Clinical Pathology.

  1. Sample preparation composite and replicate strategy case studies for assay of solid oral drug products.

    Science.gov (United States)

    Nickerson, Beverly; Harrington, Brent; Li, Fasheng; Guo, Michele Xuemei

    2017-11-30

    Drug product assay is one of several tests required for new drug products to ensure the quality of the product at release and throughout the life cycle of the product. Drug product assay testing is typically performed by preparing a composite sample of multiple dosage units to obtain an assay value representative of the batch. In some cases replicate composite samples may be prepared and the reportable assay value is the average value of all the replicates. In previously published work by Harrington et al. (2014) [5], a sample preparation composite and replicate strategy for assay was developed to provide a systematic approach which accounts for variability due to the analytical method and dosage form with a standard error of the potency assay criteria based on compendia and regulatory requirements. In this work, this sample preparation composite and replicate strategy for assay is applied to several case studies to demonstrate the utility of this approach and its application at various stages of pharmaceutical drug product development. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Sequence Optimized Real-Time RT-PCR Assay for Detection of Crimean-Congo Hemorrhagic Fever Virus

    Science.gov (United States)

    2017-03-21

    on the linear range of the curve 129 using a nonlinear regression analysis. A two-way ANOVA with Sidak’s multiple comparisons test was 130 done to...currently fielded CCHFV assay. 31 TR-17-094 DISTRIBUTION STATEMENT A: Approved for public release; distribution is unlimited. 2 Introduction 32...analyses were performed using GraphPrism 6 (GraphPad Software, San Diego, CA). 128 Assay linearity based on the preliminary LOD was determined based

  3. A radiochemical assay for biotin in biological materials

    International Nuclear Information System (INIS)

    Hood, R.L.

    1975-01-01

    A radiochemical assay for biotin is described. The assay was sensitive to one nanogram and simple enough for routine biotin analyses. The assay yielded results which were comparable to those obtained from a microbiological assay using Lactobacillus plantarum. (author)

  4. Radioreceptor assay: theory and applications to pharmacology

    International Nuclear Information System (INIS)

    Perret, G.; Simon, P.

    1984-01-01

    The aim of the first part of this work is to present the theory of the radioreceptor assay and to compare it to the other techniques of radioanalysis (radioimmunoassay, competitive protein binding assays). The technology of the radioreceptor assay is then presented and its components (preparation of the receptors, radioligand, incubation medium) are described. The analytical characteristics of the radioreceptor assay (specificity, sensitivity, reproductibility, accuracy) and the pharmacological significance of the results are discussed. The second part is devoted to the description of the radioreceptor assays of some pharmacological classes (neuroleptics, tricyclic antidepressants, benzodiazepines, β-blockers, anticholinergic drugs) and to their use in therapeutic drug monitoring. In conclusion, by their nature, radioreceptor assays are highly sensitive, reliable, precise, accurate and simple to perform. Their chief disadvantage relates to specificity, since any substance having an appreciable affinity to the receptor site will displace the specifically bound radioligand. Paradoxically in some cases, this lack of specificity may be advantageous in that it allows for the detection of not only the apparent compound but of active metabolites and endogenous receptor agonists as well and in that radioreceptors assays can be devised for a whole pharmacological class and not only for one drug as it is the case for classical physico-chemical techniques. For all these reasons future of radioreceptor assay in pharmacology appears promising [fr

  5. Assessing sediment contamination using six toxicity assays

    Directory of Open Access Journals (Sweden)

    Allen G. BURTON Jr.

    2001-08-01

    Full Text Available An evaluation of sediment toxicity at Lake Orta, Italy was conducted to compare a toxicity test battery of 6 assays and to evaluate the extent of sediment contamination at various sediment depths. Lake Orta received excessive loadings of copper and ammonia during the 1900’s until a large remediation effort was conducted in 1989-90 using lime addition. Since that time, the lake has shown signs of a steady recovery of biological communities. The study results showed acute toxicity still exists in sediments at a depth of 5 cm and greater. Assays that detected the highest levels of toxicity were two whole sediment exposures (7 d using Hyalella azteca and Ceriodaphnia dubia. The MicrotoxR assay using pore water was the third most sensitive assay. The Thamnotox, Rototox, Microtox solid phase, and Seed Germination-Root Elongation (pore and solid phase assays showed occasional to no toxicity. Based on similarity of responses and assay sensitivity, the two most useful assays were the C. dubia (or H. azteca and Microtox pore water. These assays were effective at describing sediment toxicity in a weight-of-evidence approach.

  6. A Continuous, Fluorogenic Sirtuin 2 Deacylase Assay

    DEFF Research Database (Denmark)

    Galleano, Iacopo; Schiedel, Matthias; Jung, Manfred

    2016-01-01

    and kinetic insight regarding sirtuin inhibitors, it is important to have access to efficient assays. In this work, we report readily synthesized fluorogenic substrates enabling enzyme-economical evaluation of SIRT2 inhibitors in a continuous assay format as well as evaluation of the properties of SIRT2...

  7. 21 CFR 864.7525 - Heparin assay.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification. A...

  8. Detection of IFN-γ Secretion by T Cells Collected Before and After Successful Treatment of Early Lyme Disease.

    Science.gov (United States)

    Callister, Steven M; Jobe, Dean A; Stuparic-Stancic, Aleksandra; Miyamasu, Misato; Boyle, Jeff; Dattwyler, Raymond J; Arnaboldi, Paul M

    2016-05-15

    Current serodiagnostics for Lyme disease lack sensitivity during early disease, and cannot determine treatment response. We evaluated an assay based on QuantiFERON technology utilizing peptide antigens derived from Borrelia burgdorferi to stimulate interferon-gamma (IFN-γ) release as an alternative to serodiagnosis for the laboratory detection of Lyme disease. Blood was obtained from patients with erythema migrans before (n = 29) and 2 months after (n = 27) antibiotic therapy. IFN-γ release was measured by enzyme-linked immunosorbent assay (ELISA) following overnight stimulation of whole blood with the peptide antigens, and compared to the results of standard serological assays (C6, ELISA, and Western blot). IFN-γ release was observed in pretreatment blood of 20 of 29 (69%) patients with Lyme disease. Following antibiotic treatment, IFN-γ was significantly reduced (P = .0002), and was detectable in only 4 of 20 (20%) initially positive patients. By contrast, anti-C6 antibodies were detected in pretreatment sera from 17 of 29 (59%) subjects, whereas only 5 of 29 (17%) patients had positive Western blot seroreactivity. Antibody responses persisted and expanded following treatment. Our findings suggest that measurement of IFN-γ after incubating blood with Borrelia antigens could be useful in the laboratory diagnosis of early Lyme disease. Also, after antibiotic treatment, this response appears to be short lived. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  9. Release plan for Big Pete

    International Nuclear Information System (INIS)

    Edwards, T.A.

    1996-11-01

    This release plan is to provide instructions for the Radiological Control Technician (RCT) to conduct surveys for the unconditional release of ''Big Pete,'' which was used in the removal of ''Spacers'' from the N-Reactor. Prior to performing surveys on the rear end portion of ''Big Pete,'' it shall be cleaned (i.e., free of oil, grease, caked soil, heavy dust). If no contamination is found, the vehicle may be released with the permission of the area RCT Supervisor. If contamination is found by any of the surveys, contact the cognizant Radiological Engineer for decontamination instructions

  10. Commercial SNF Accident Release Fractions

    Energy Technology Data Exchange (ETDEWEB)

    J. Schulz

    2004-11-05

    The purpose of this analysis is to specify and document the total and respirable fractions for radioactive materials that could be potentially released from an accident at the repository involving commercial spent nuclear fuel (SNF) in a dry environment. The total and respirable release fractions are used to support the preclosure licensing basis for the repository. The total release fraction is defined as the fraction of total commercial SNF assembly inventory, typically expressed as an activity inventory (e.g., curies), of a given radionuclide that is released to the environment from a waste form. Radionuclides are released from the inside of breached fuel rods (or pins) and from the detachment of radioactive material (crud) from the outside surfaces of fuel rods and other components of fuel assemblies. The total release fraction accounts for several mechanisms that tend to retain, retard, or diminish the amount of radionuclides that are available for transport to dose receptors or otherwise can be shown to reduce exposure of receptors to radiological releases. The total release fraction includes a fraction of airborne material that is respirable and could result in inhalation doses; this subset of the total release fraction is referred to as the respirable release fraction. Accidents may involve waste forms characterized as: (1) bare unconfined intact fuel assemblies, (2) confined intact fuel assemblies, or (3) canistered failed commercial SNF. Confined intact commercial SNF assemblies at the repository are contained in shipping casks, canisters, or waste packages. Four categories of failed commercial SNF are identified: (1) mechanically and cladding-penetration damaged commercial SNF, (2) consolidated/reconstituted assemblies, (3) fuel rods, pieces, and debris, and (4) nonfuel components. It is assumed that failed commercial SNF is placed into waste packages with a mesh screen at each end (CRWMS M&O 1999). In contrast to bare unconfined fuel assemblies, the

  11. Commercial SNF Accident Release Fractions

    International Nuclear Information System (INIS)

    Schulz, J.

    2004-01-01

    The purpose of this analysis is to specify and document the total and respirable fractions for radioactive materials that could be potentially released from an accident at the repository involving commercial spent nuclear fuel (SNF) in a dry environment. The total and respirable release fractions are used to support the preclosure licensing basis for the repository. The total release fraction is defined as the fraction of total commercial SNF assembly inventory, typically expressed as an activity inventory (e.g., curies), of a given radionuclide that is released to the environment from a waste form. Radionuclides are released from the inside of breached fuel rods (or pins) and from the detachment of radioactive material (crud) from the outside surfaces of fuel rods and other components of fuel assemblies. The total release fraction accounts for several mechanisms that tend to retain, retard, or diminish the amount of radionuclides that are available for transport to dose receptors or otherwise can be shown to reduce exposure of receptors to radiological releases. The total release fraction includes a fraction of airborne material that is respirable and could result in inhalation doses; this subset of the total release fraction is referred to as the respirable release fraction. Accidents may involve waste forms characterized as: (1) bare unconfined intact fuel assemblies, (2) confined intact fuel assemblies, or (3) canistered failed commercial SNF. Confined intact commercial SNF assemblies at the repository are contained in shipping casks, canisters, or waste packages. Four categories of failed commercial SNF are identified: (1) mechanically and cladding-penetration damaged commercial SNF, (2) consolidated/reconstituted assemblies, (3) fuel rods, pieces, and debris, and (4) nonfuel components. It is assumed that failed commercial SNF is placed into waste packages with a mesh screen at each end (CRWMS M andO 1999). In contrast to bare unconfined fuel assemblies, the

  12. Highly Efficient Thermoresponsive Nanocomposite for Controlled Release Applications

    KAUST Repository

    Yassine, Omar

    2016-06-23

    Highly efficient magnetic release from nanocomposite microparticles is shown, which are made of Poly (N-isopropylacrylamide) hydrogel with embedded iron nanowires. A simple microfluidic technique was adopted to fabricate the microparticles with a high control of the nanowire concentration and in a relatively short time compared to chemical synthesis methods. The thermoresponsive microparticles were used for the remotely triggered release of Rhodamine (B). With a magnetic field of only 1 mT and 20 kHz a drug release of 6.5% and 70% was achieved in the continuous and pulsatile modes, respectively. Those release values are similar to the ones commonly obtained using superparamagnetic beads but accomplished with a magnetic field of five orders of magnitude lower power. The high efficiency is a result of the high remanent magnetization of the nanowires, which produce a large torque when exposed to a magnetic field. This causes the nanowires to vibrate, resulting in friction losses and heating. For comparison, microparticles with superparamagnetic beads were also fabricated and tested; while those worked at 73 mT and 600 kHz, no release was observed at the low field conditions. Cytotoxicity assays showed similar and high cell viability for microparticles with nanowires and beads.

  13. Highly Efficient Thermoresponsive Nanocomposite for Controlled Release Applications

    KAUST Repository

    Yassine, Omar; Zaher, Amir; Li, Erqiang; Alfadhel, Ahmed; Perez, Jose E.; Kavaldzhiev, Mincho; Contreras, Maria F.; Thoroddsen, Sigurdur T; Khashab, Niveen M.; Kosel, Jü rgen

    2016-01-01

    Highly efficient magnetic release from nanocomposite microparticles is shown, which are made of Poly (N-isopropylacrylamide) hydrogel with embedded iron nanowires. A simple microfluidic technique was adopted to fabricate the microparticles with a high control of the nanowire concentration and in a relatively short time compared to chemical synthesis methods. The thermoresponsive microparticles were used for the remotely triggered release of Rhodamine (B). With a magnetic field of only 1 mT and 20 kHz a drug release of 6.5% and 70% was achieved in the continuous and pulsatile modes, respectively. Those release values are similar to the ones commonly obtained using superparamagnetic beads but accomplished with a magnetic field of five orders of magnitude lower power. The high efficiency is a result of the high remanent magnetization of the nanowires, which produce a large torque when exposed to a magnetic field. This causes the nanowires to vibrate, resulting in friction losses and heating. For comparison, microparticles with superparamagnetic beads were also fabricated and tested; while those worked at 73 mT and 600 kHz, no release was observed at the low field conditions. Cytotoxicity assays showed similar and high cell viability for microparticles with nanowires and beads.

  14. Cholecystokinin receptors: disparity between phosphoinositide breakdown and amylase releasing activity of CCK analogues in pancreas

    International Nuclear Information System (INIS)

    Lin, C.W.; Grant, D.; Bianchi, B.; Miller, T.; Witte, D.; Shue, Y.K.; Nadzan, A.

    1986-01-01

    Cholecystokinin (CCK) peptides are a family of hormones which also occur in brain. In pancreas CCK stimulates the release of amylase, a process that is dependent on the mobilization of intracellular Ca 2+ . Recent evidence suggests that inositol 1,4,5-trisphosphate, the breakdown product of phosphatidylinositol 4,5-bisphosphate, is responsible for the rise in intracellular Ca 2+ . Their laboratory has developed assays to study synthetic CCK analogues using radioligand binding, PI breakdown and amylase release. They have shown that there are good correlations among these three assay systems for the carboxy terminal fragments of CCK 8 . Recently, they have discovered synthetic analogues of CCK 4 that are full agonists in amylase release but are ineffective in causing PI breakdown. In particular, A-61576, Boc-5-amino-2-indolemethylene-pent-2-ene-1-oyl-Leu-Asp-Phe-NH 2 , is a full agonist in the amylase releasing assay, but is devoid of PI stimulating activity. A-61576 completely reverses the stimulation of PI response induced by CCK 8 , indicative of an antagonist. Since a mechanism other than the PI breakdown is responsible for amylase release by A-61576, they suggest that separate receptors are responsible for PI breakdown and amylase release

  15. Sensitive microplate assay for the detection of proteolytic enzymes using radiolabeled gelatin

    International Nuclear Information System (INIS)

    Robertson, B.D.; Kwan-Lim, G.E.; Maizels, R.M.

    1988-01-01

    A sensitive, microplate assay is described for the detection of a wide range of proteolytic enzymes, using radio-iodine-labeled gelatin as substrate. The technique uses the Bolton-Hunter reagent to label the substrate, which is then coated onto the wells of polyvinyl chloride microtiter plates. By measuring the radioactivity released the assay is able to detect elastase, trypsin, and collagenase in concentrations of 1 ng/ml or less, while the microtiter format permits multiple sample handling and minimizes sample volumes required for analysis

  16. Comparison of Batch Assay and Random Assay Using Automatic Dispenser in Radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Moon, Seung Hwan; Jang, Su Jin; Kang, Ji Yeon; Lee, Dong Soo; Chung, June Key; Lee, Myung Chul [Seoul Metropolitan Government Seoul National University Boramae Medical Center, Seoul (Korea, Republic of); Lee, Ho Young; Shin, Sun Young; Min, Gyeong Sun; Lee, Hyun Joo [Seoul National University college of Medicine, Seoul (Korea, Republic of)

    2009-08-15

    Radioimmunoassay (RIA) was usually performed by the batch assay. To improve the efficiency of RIA without increase of the cost and time, random assay could be a choice. We investigated the possibility of the random assay using automatic dispenser by assessing the agreement between batch assay and random assay. The experiments were performed with four items; Triiodothyronine (T3), free thyroxine (fT4), Prostate specific antigen (PSA), Carcinoembryonic antigen (CEA). In each item, the sera of twenty patients, the standard, and the control samples were used. The measurements were done 4 times with 3 hour time intervals by random assay and batch assay. The coefficient of variation (CV) of the standard samples and patients' data in T3, fT4, PSA, and CEA were assessed. ICC (Intraclass correlation coefficient) and coefficient of correlation were measured to assessing the agreement between two methods. The CVs (%) of T3, fT4, PSA, and CEA measured by batch assay were 3.2+-1.7%, 3.9+-2.1%, 7.1+-6.2%, 11.2+-7.2%. The CVs by random assay were 2.1+-1.7%, 4.8+-3.1%, 3.6+-4.8%, and 7.4+-6.2%. The ICC between the batch assay and random assay were 0.9968 (T3), 0.9973 (fT4), 0.9996 (PSA), and 0.9901 (CEA). The coefficient of correlation between the batch assay and random assay were 0.9924(T3), 0.9974 (fT4), 0.9994 (PSA), and 0.9989 (CEA) (p<0.05). The results of random assay showed strong agreement with the batch assay in a day. These results suggest that random assay using automatic dispenser could be used in radioimmunoassay

  17. Comparison of Batch Assay and Random Assay Using Automatic Dispenser in Radioimmunoassay

    International Nuclear Information System (INIS)

    Moon, Seung Hwan; Jang, Su Jin; Kang, Ji Yeon; Lee, Dong Soo; Chung, June Key; Lee, Myung Chul; Lee, Ho Young; Shin, Sun Young; Min, Gyeong Sun; Lee, Hyun Joo

    2009-01-01

    Radioimmunoassay (RIA) was usually performed by the batch assay. To improve the efficiency of RIA without increase of the cost and time, random assay could be a choice. We investigated the possibility of the random assay using automatic dispenser by assessing the agreement between batch assay and random assay. The experiments were performed with four items; Triiodothyronine (T3), free thyroxine (fT4), Prostate specific antigen (PSA), Carcinoembryonic antigen (CEA). In each item, the sera of twenty patients, the standard, and the control samples were used. The measurements were done 4 times with 3 hour time intervals by random assay and batch assay. The coefficient of variation (CV) of the standard samples and patients' data in T3, fT4, PSA, and CEA were assessed. ICC (Intraclass correlation coefficient) and coefficient of correlation were measured to assessing the agreement between two methods. The CVs (%) of T3, fT4, PSA, and CEA measured by batch assay were 3.2±1.7%, 3.9±2.1%, 7.1±6.2%, 11.2±7.2%. The CVs by random assay were 2.1±1.7%, 4.8±3.1%, 3.6±4.8%, and 7.4±6.2%. The ICC between the batch assay and random assay were 0.9968 (T3), 0.9973 (fT4), 0.9996 (PSA), and 0.9901 (CEA). The coefficient of correlation between the batch assay and random assay were 0.9924(T3), 0.9974 (fT4), 0.9994 (PSA), and 0.9989 (CEA) (p<0.05). The results of random assay showed strong agreement with the batch assay in a day. These results suggest that random assay using automatic dispenser could be used in radioimmunoassay

  18. Biomonitoring of genotoxic risk in radar facility workers: comparison of the comet assay with micronucleus assay and chromatid breakage assay

    International Nuclear Information System (INIS)

    Garaj-Vrhovac, V.; Kopjar, N.

    2003-01-01

    Genotoxic risks of occupational exposure in a radar facility were evaluated by using alkaline comet assay, micronucleus assay and chromatid breakage assay on peripheral blood leukocytes in exposed subjects and corresponding controls. Results show that occupational exposure to microwave radiation correlates with an increase of genome damage in somatic cells. The levels of DNA damage in exposed subjects determined by using alkaline comet assay were increased compared to control and showed interindividual variations. Incidence of micronuclei was also significantly increased compared to baseline control values. After short exposure of cultured lymphocytes to bleomycin, cells of occupationally exposed subjects responded with high numbers of chromatid breaks. Although the level of chromosome damage generated by bleomycin varied greatly between individuals, in exposed subjects a significantly elevated number of chromatid breaks was observed. Our results support data reported in literature indicating that microwave radiation represents a potential DNA-damaging hazard. Alkaline comet assay is confirmed as a sensitive and highly reproducible technique for detection of primary DNA damage inflicted in somatic cells. Micronucleus assay was confirmed as reliable bio-markers of effect and chromatid breakage assay as sensitive bio-marker of individual cancer susceptibility. The results obtained also confirm the necessity to improve measures and to perform accurate health surveillance of individuals occupationally exposed to microwave radiation

  19. Immunological assays in the diagnosis of tuberculosis infection at the beginning of 21st century

    Directory of Open Access Journals (Sweden)

    Kašiković-Lečić Svetlana

    2010-01-01

    Full Text Available A subject infected by tubercle bacilli may immediately develop the disease (primary tuberculosis, witch occurs in a few number of infected persons, or may overcome the primary infection (a latent carrier of the infection. Nowadays there are about two milliard people with a latent tubercular infection. About 10% of them will develop active tuberculosis during their lifetime. Detection of latent tuberculosis and its treatment decreases the risk of developing the active form of the disease. Until recently, the tuberculin test was a single screening method to identify the subjects with tubercular infection. In recent years, some novel in vitro tests to diagnose the tubercular infection have been designed. These tests measure the cell-mediated immune response quantifying the emission of interferon gamma by T cells in response to stimulation by Mycobacterium tuberculosis specific antigens. These tests are more sensitive and specific than the tuberculin skin test, but they are just an accessory tool, i.e. a link in the diagnostic chain of the tubercular infection.

  20. Birth control - slow release methods

    Science.gov (United States)

    Contraception - slow-release hormonal methods; Progestin implants; Progestin injections; Skin patch; Vaginal ring ... might want to consider a different birth control method. SKIN PATCH The skin patch is placed on ...

  1. DEVELOPMENT OF SUSTAINED RELEASE TABLETS ...

    African Journals Online (AJOL)

    2013-12-31

    Dec 31, 2013 ... The SR dosage forms that release drugs pH independently in .... were determined; Post compression parameters such as weight variation test, hardness, ... Based on the ICH guidelines 12, the stability studies were carried out ...

  2. Exosomes released from breast cancer carcinomas stimulate cell movement.

    Directory of Open Access Journals (Sweden)

    Dinari A Harris

    Full Text Available For metastasis to occur cells must communicate with to their local environment to initiate growth and invasion. Exosomes have emerged as an important mediator of cell-to-cell signalling through the transfer of molecules such as mRNAs, microRNAs, and proteins between cells. Exosomes have been proposed to act as regulators of cancer progression. Here, we study the effect of exosomes on cell migration, an important step in metastasis. We performed cell migration assays, endocytosis assays, and exosome proteomic profiling on exosomes released from three breast cancer cell lines that model progressive stages of metastasis. Results from these experiments suggest: (1 exosomes promote cell migration and (2 the signal is stronger from exosomes isolated from cells with higher metastatic potentials; (3 exosomes are endocytosed at the same rate regardless of the cell type; (4 exosomes released from cells show differential enrichment of proteins with unique protein signatures of both identity and abundance. We conclude that breast cancer cells of increasing metastatic potential secrete exosomes with distinct protein signatures that proportionally increase cell movement and suggest that released exosomes could play an active role in metastasis.

  3. PCDD/PCDF release inventories

    Energy Technology Data Exchange (ETDEWEB)

    Fiedler, H. [UNEP Chemicals, Chatelaine (Switzerland)

    2004-09-15

    The Stockholm Convention on Persistent Organic Pollutants (POPs) entered into force on 17 May 2004 with 50 Parties. In May 2004, 59 countries had ratified or acceded the Convention. The objective of the Convention is ''to protect human health and the environment from persistent organic pollutants''. For intentionally produced POPs, e.g., pesticides and industrial chemicals such as hexachlorobenzene and polychlorinated biphenyls, this will be achieved by stop of production and use. For unintentionally generated POPs, such as polychlorinated dibenzo-pdioxins (PCDD) and polychlorinated dibenzofurans (PCDF), measures have to be taken to ''reduce the total releases derived from anthropogenic sources''; the final goal is ultimate elimination, where feasible. Under the Convention, Parties have to establish and maintain release inventories to prove the continuous release reduction. Since many countries do not have the technical and financial capacity to measure all releases from all potential PCDD/PCDF sources, UNEP Chemicals has developed the ''Standardized Toolkit for the Identification of Quantification of Dioxin and Furan Releases'' (''Toolkit'' for short), a methodology to estimate annual releases from a number of sources. With this methodology, annual releases can be estimated by multiplying process-specific default emission factors provided in the Toolkit with national activity data. At the seventh session of the Intergovernmental Negotiating Committee, the Toolkit was recommended to be used by countries when reporting national release data to the Conference of the Parties. The Toolkit is especially used by developing countries and countries with economies in transition where no measured data are available. Results from Uruguay, Thailand, Jordan, Philippines, and Brunei Darussalam have been published.

  4. Development of a solid-phase assay for measurement of proteolytic enzyme activity

    International Nuclear Information System (INIS)

    Varani, J.; Johnson, K.; Kaplan, J.

    1980-01-01

    A solid-phase, plate assay was developed for the measurement of proteolytic enzyme activity. In this assay procedure, radiolabeled substrates were dried onto the surface of microtiter wells. Following drying, the wells were washed two times with saline to remove the nonadherent substrate. When proteolytic enzymes were added to the wells, protein hydrolysis occurred, releasing radioactivity into the supernatant fluid. The amount of protein hydrolysis that occurred was reflected by the amount of radioactivity in the supernatant fluid. When 125 I-hemoglobin was used as the substrate, it was as susceptible to hydrolysis by trypsin in the solid-phase assay as it was in solution in a standard assay procedure. Protease activity from a variety of sources (including from viable cells as well as from extracellular sources) were also able to hydrolyze the hemoglobin on the plate. 125 I-Labeled serum albumen, fibrinogen, and rat pulmonary basement membrane were also susceptible to hydrolysis by trypsin in the solid phase. When [ 14 C]elastin was dried onto the plate, it behaved in a similar manner to elastin in solution. It was resistant to hydrolysis by nonspecific proteases such as trypsin and chymotrypsin but was highly susceptible to hydrolysis by elastase. The solid-phase plate assay has several features which recommended it for routine use. It is as sensitive as standard tube assays (and much more sensitive than routinely used colormetric assays). It is quick and convenient; there are no precipitation, centrifugation, or filtration steps. In addition, very small volumes of radioactive wastes are generated. Another advantage of the solid-phase plate assay is the resistance of the dried substrates to spontaneous breakdown and to microbial contamination. Finally, this assay is suitable for use with viable cells as well as for extracellular proteases

  5. A high-throughput assay of NK cell activity in whole blood and its clinical application

    International Nuclear Information System (INIS)

    Lee, Saet-byul; Cha, Junhoe; Kim, Im-kyung; Yoon, Joo Chun; Lee, Hyo Joon; Park, Sang Woo; Cho, Sunjung; Youn, Dong-Ye; Lee, Heyja; Lee, Choong Hwan; Lee, Jae Myun; Lee, Kang Young; Kim, Jongsun

    2014-01-01

    Graphical abstract: - Highlights: • We demonstrated a simple assay of NK cell activity from whole blood. • The measurement of secreted IFN-γ from NK cell enables high-throughput screening. • The NKA assay was validated by clinical results of colorectal cancer patients. - Abstract: Natural killer (NK) cells are lymphocytes of the innate immune system and have the ability to kill tumor cells and virus-infected cells without prior sensitization. Malignant tumors and viruses have developed, however, strategies to suppress NK cells to escape from their responses. Thus, the evaluation of NK cell activity (NKA) could be invaluable to estimate the status and the outcome of cancers, viral infections, and immune-mediated diseases. Established methods that measure NKA, such as 51 Cr release assay and CD107a degranulation assay, may be used to determine NK cell function, but they are complicated and time-consuming because they require isolation of peripheral blood mononuclear cells (PBMC) or NK cells. In some cases these assays require hazardous material such as radioactive isotopes. To overcome these difficulties, we developed a simple assay that uses whole blood instead of PBMC or isolated NK cells. This novel assay is suitable for high-throughput screening and the monitoring of diseases, because it employs serum of ex vivo stimulated whole blood to detect interferon (IFN)-γ secreted from NK cells as an indicator of NKA. After the stimulation of NK cells, the determination of IFNγ concentration in serum samples by enzyme-linked immunosorbent assay (ELISA) provided a swift, uncomplicated, and high-throughput assay of NKA ex vivo. The NKA results microsatellite stable (MSS) colorectal cancer patients was showed significantly lower NKA, 263.6 ± 54.5 pg/mL compared with healthy subjects, 867.5 ± 50.2 pg/mL (p value <0.0001). Therefore, the NKA could be utilized as a supportive diagnostic marker for microsatellite stable (MSS) colorectal cancer

  6. A high-throughput assay of NK cell activity in whole blood and its clinical application

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Saet-byul [Department of Microbiology and Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul (Korea, Republic of); Cha, Junhoe [ATGen Co. Ltd., Sungnam (Korea, Republic of); Kim, Im-kyung [Department of Surgery, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul (Korea, Republic of); Yoon, Joo Chun [Department of Microbiology, Ewha Womans University School of Medicine, Seoul (Korea, Republic of); Lee, Hyo Joon [Department of Microbiology and Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul (Korea, Republic of); Park, Sang Woo; Cho, Sunjung; Youn, Dong-Ye; Lee, Heyja; Lee, Choong Hwan [ATGen Co. Ltd., Sungnam (Korea, Republic of); Lee, Jae Myun [Department of Microbiology and Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul (Korea, Republic of); Lee, Kang Young, E-mail: kylee117@yuhs.ac [Department of Surgery, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul (Korea, Republic of); Kim, Jongsun, E-mail: jkim63@yuhs.ac [Department of Microbiology and Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul (Korea, Republic of)

    2014-03-14

    Graphical abstract: - Highlights: • We demonstrated a simple assay of NK cell activity from whole blood. • The measurement of secreted IFN-γ from NK cell enables high-throughput screening. • The NKA assay was validated by clinical results of colorectal cancer patients. - Abstract: Natural killer (NK) cells are lymphocytes of the innate immune system and have the ability to kill tumor cells and virus-infected cells without prior sensitization. Malignant tumors and viruses have developed, however, strategies to suppress NK cells to escape from their responses. Thus, the evaluation of NK cell activity (NKA) could be invaluable to estimate the status and the outcome of cancers, viral infections, and immune-mediated diseases. Established methods that measure NKA, such as {sup 51}Cr release assay and CD107a degranulation assay, may be used to determine NK cell function, but they are complicated and time-consuming because they require isolation of peripheral blood mononuclear cells (PBMC) or NK cells. In some cases these assays require hazardous material such as radioactive isotopes. To overcome these difficulties, we developed a simple assay that uses whole blood instead of PBMC or isolated NK cells. This novel assay is suitable for high-throughput screening and the monitoring of diseases, because it employs serum of ex vivo stimulated whole blood to detect interferon (IFN)-γ secreted from NK cells as an indicator of NKA. After the stimulation of NK cells, the determination of IFNγ concentration in serum samples by enzyme-linked immunosorbent assay (ELISA) provided a swift, uncomplicated, and high-throughput assay of NKA ex vivo. The NKA results microsatellite stable (MSS) colorectal cancer patients was showed significantly lower NKA, 263.6 ± 54.5 pg/mL compared with healthy subjects, 867.5 ± 50.2 pg/mL (p value <0.0001). Therefore, the NKA could be utilized as a supportive diagnostic marker for microsatellite stable (MSS) colorectal cancer.

  7. Nano-immunosafety: issues in assay validation

    International Nuclear Information System (INIS)

    Boraschi, Diana; Italiani, Paola; Oostingh, Gertie J; Duschl, Albert; Casals, Eudald; Puntes, Victor F; Nelissen, Inge

    2011-01-01

    Assessing the safety of engineered nanomaterials for human health must include a thorough evaluation of their effects on the immune system, which is responsible for defending the integrity of our body from damage and disease. An array of robust and representative assays should be set up and validated, which could be predictive of the effects of nanomaterials on immune responses. In a trans-European collaborative work, in vitro assays have been developed to this end. In vitro tests have been preferred for their suitability to standardisation and easier applicability. Adapting classical assays to testing the immunotoxicological effects of nanoparticulate materials has raised a series of issues that needed to be appropriately addressed in order to ensure reliability of results. Besides the exquisitely immunological problem of selecting representative endpoints predictive of the risk of developing disease, assay results turned out to be significantly biased by artefactual interference of the nanomaterials or contaminating agents with the assay protocol. Having addressed such problems, a series of robust and representative assays have been developed that describe the effects of engineered nanoparticles on professional and non-professional human defence cells. Two of such assays are described here, one based on primary human monocytes and the other employing human lung epithelial cells transfected with a reporter gene.

  8. Radioactive wastes assay technique and equipment

    International Nuclear Information System (INIS)

    Lee, K. M.; Hong, D. S; Kim, T. K.; Bae, S. M.; Shon, J. S.; Hong, K. P.

    2004-12-01

    The waste inventory records such as the activities and radio- nuclides contained in the waste packages are to be submitted with the radioactive wastes packages for the final disposal. The nearly around 10,000 drums of waste stocked in KAERI now should be assayed for the preparation of the waste inventory records too. For the successive execution of the waste assay, the investigation into the present waste assay techniques and equipment are to be taken first. Also the installation of the waste assay equipment through the comprehensive design, manufacturing and procurement should be proceeded timely. As the characteristics of the KAERI-stocked wastes are very different from that of the nuclear power plant and those have no regular waste streams, the application of the in-direct waste assay method using the scaling factors are not effective for the KAERI-generated wastes. Considering for the versal conveniency including the accuracy over the wide range of waste forms and the combination of assay time and sensitivity, the TGS(Tomographic Gamma Scanner) is appropriate as for the KAERI -generated radioactive waste assay equipment

  9. A multiwell format assay for heparanase.

    Science.gov (United States)

    Behzad, Farhad; Brenchley, Paul E C

    2003-09-15

    This assay employs a biotinylated heparan sulfate glycosaminoglycan (HSGAG) substrate that is covalently linked to the surface of 96-well immunoassay plates. The ratio of biotin:HSGAG and the coating concentration of substrate bound to the wells have been optimized and allow removal of biotin HSGAG within 60 min of incubation at 37 degrees C in assay buffer with a standard dilution of bacterial heparitinase or platelet heparanase. Loss of biotin signal from the well surface is detected on incubation with peroxidase-streptavidin followed by color development using 3,3',5,5'-tetramethylbenzidine as the peroxidase substrate. The new assay allows specific detection of heparanase activity in multiple samples in a total time of 3 h including a 1-h substrate digestion step and is a significant improvement with regard to sensitivity, specificity, and ease of handling of multiple samples compared to other described assays. Heparanase specifically degrades the biotinylated HSGAG substrate, when used with an optimized assay buffer. A range of enzymes including collagenase, trypsin, plasmin, pepsin, chondroitinases, hyaluronidase, and neuraminidase show no effect on the substrate under optimized assay conditions. The covalent linkage of the substrate to the well prevents leaching of substrate and allows preparation and long-term storage of substrate-coated plates. The assay can be used to detect heparanase levels in clinical samples and cell culture supernatants and is ideal as a screening method for antagonists of enzyme activity.

  10. Hydraulic running and release tool with mechanical emergency release

    International Nuclear Information System (INIS)

    Baker, S.F.

    1991-01-01

    This patent describes a setting tool for connection in a well string to position a tubular member in a well bore. It comprises: a mandrel adapted to be connected to the well string; an outer sleeve surrounding the mandrel and releasably secured thereto; a latch nut releasably connected to the outer sleeve; piston means sealingly engaging the mandrel; shear means releasably securing the piston to the latch nut to maintain the latch nut releasably connected to the tubular member; the mandrel having port means for conducting fluid pressure from the well string to release the piston means from and the latch nut; cooperating engageable surfaces on the piston and latch nut to reengage them together after the piston moves a predetermined longitudinal distance relative to the latch nut; and additional cooperating engageable surfaces on the latch nut and the outer sleeve which are engageable when the piston and engaged latch nut are moved a predetermined additional longitudinal distance by fluid pressure to secure the engaged piston and latch nut with the outer sleeve for retrieval along with the mandrel from the well bore

  11. Modulation of cytokine release by differentiated CACO-2 cells in a compartmentalized coculture model with mononuclear leucocytes and nonpathogenic bacteria

    DEFF Research Database (Denmark)

    Parlesak, Alexandr; Haller, D.; Brinz, S.

    2004-01-01

    To further investigate the interaction between human mononuclear leucocytes [peripheral blood mononuclear cells (PBMC)] and enterocytes, the effect of a confluent layer of differentiated CACO-2 cells on cytokine kinetics during challenge with bacteria in a compartmentalized coculture model...... cells when leucocytes were stimulated directly with bacteria. This suppression was not paralleled by changes in the production of IL-10, IL-6 and transforming growth factor (TGF)-beta. When the bacteria were applied apically to the CACO-2 cell layer, the production of TNF-alpha, IL-12, IL-1beta, IL-8......, IL-6, IL-10, TGF-beta and interferon-gamma was pronouncedly lower as compared to the bacterial stimulation of leucocytes beneath the CACO-2 cells. In the latter experiments, IL-6, IL-8 and TNF-alpha were the cytokines being mostly induced by apical addition of E. coli. Quantitative mRNA expression...

  12. Nondestructive assay measurements applied to reprocessing plants

    International Nuclear Information System (INIS)

    Ruhter, Wayne D.; Lee, R. Stephen; Ottmar, Herbert; Guardini, Sergio

    1999-01-01

    Nondestructive assay for reprocessing plants relies on passive gamma-ray spectrometry for plutonium isotopic and plutonium mass values of medium-to-low-density samples and holdup deposits; on active x-ray fluorescence and densitometry techniques for uranium and plutonium concentrations in solutions; on calorimetry for plutonium mass in product; and passive neutron techniques for plutonium mass in spent fuel, product, and waste. This paper will describe the radiation-based nondestructive assay techniques used to perform materials accounting measurements. The paper will also discuss nondestructive assay measurements used in inspections of reprocessing plants [ru

  13. Thermometric enzyme linked immunosorbent assay: TELISA.

    Science.gov (United States)

    Mattiasson, B; Borrebaeck, C; Sanfridson, B; Mosbach, K

    1977-08-11

    A new method, thermometric enzyme linked immunosorbent assay (TELISA), for the assay of endogenous and exogenous compounds in biological fluids is described. It is based on the previously described enzyme linked immunosorbent assay technique, ELISA, but utilizes enzymic heat formation which is measured in an enzyme thermistor unit. In the model system studied determination of human serum albumin down to a concentration of 10(-10) M (5 ng/ml) was achieved, with both normal and catalase labelled human serum albumin competing for the binding sites on the immunosorbent, which was rabbit antihuman serum albumin immobilized onto Sepharose CL-4B.

  14. Comparison of multiple assays for detecting human antibodies directed against antigens on normal and malignant tissue culture cells

    International Nuclear Information System (INIS)

    Rosenberg, S.A.; Schwarz, S.; Anding, H.; Hyatt, C.; Williams, G.M.; Johns Hopkins Univ., Baltimore, Md.

    1977-01-01

    Four separate assays of human antibody reactivity to four separate normal and malignant human tissue culture cells lines from two patients have been evaluated using a single highly-reactive allogeneic serum. The visual end-point cytolysis assay and the chromium-51 release assay were equally sensitive in measuring complement mediated antibody cytotoxicity and both were far more sensitive than a trypan blue dye exclusion assay. The assay of antibody reactivity by hemadsorption technique was about 10 times more sensitive than any of the cytotoxicity assays. This latter assay measures only IgG antibody however. These assays showed that cell lines from different patients may differ greatly in 'reactivity' to an allogeneic serum and emphasized the importance of utilizing tumor and normal cells from the same patient when using tissue culture cells to search for tumor specific reactivity. These observations emphasize the importance of utilizing multiple assays against paired normal and malignant cells from the same patient to be certain of the specificity and magnitude of the measured antibody

  15. Lipase assay in soils by copper soap colorimetry.

    Science.gov (United States)

    Saisuburamaniyan, N; Krithika, L; Dileena, K P; Sivasubramanian, S; Puvanakrishnan, R

    2004-07-01

    A simple and sensitive method for the estimation of lipase activity in soils is reported. In this method, 50mg of soil is incubated with emulsified substrate, the fatty acids liberated are treated with cupric acetate-pyridine reagent, and the color developed is measured at 715 nm. Use of olive oil in this protocol leads to an estimation of true lipase activity in soils. The problem of released fatty acids getting adsorbed onto the soil colloids is obviated by the use of isooctane, and separate standards for different soils need not be developed. Among the various surfactants used for emulsification, polyvinyl alcohol is found to be the most effective. Incubation time of 20 min, soil concentration of 50 mg, pH 6.5, and incubation temperature of 37 degrees C were found to be the most suitable conditions for this assay. During the process of enrichment of the soils with oil, interference by the added oil is avoided by the maintenance of a suitable control, wherein 50 mg of soil is added after stopping the reaction. This assay is sensitive and it could be adopted to screen for lipase producers from enriched soils and oil-contaminated soils before resorting to isolation of the microbes by classical screening methods.

  16. Tests for oil/dispersant toxicity: In situ laboratory assays

    International Nuclear Information System (INIS)

    Wright, D.A.; Coelho, G.M.; Aurand, D.V.

    1995-01-01

    As part of its readiness program in oil spill response, the Marine Pollution Control Unit (MPCU), Department of Transport, U.K. conducts annual field trials in the North Sea, approximately 30 nautical miles from the southeast coast of England. The trials take the form of controlled releases of crude oil or Medium Fuel/Gas Oil mix (MFO), with and without the application of Corexit 9527 dispersant. In 1994 and 1995 the authors conducted a series of in situ toxicity bioassays in association with these spills with included 48h LC50 tests for turbot (Scophthalmus maximus) and oyster (Crassostrea gigas) larvae, a 48 h oyster (C. gigas) embryonic development test and two full life-cycle assays using the copepods Acartia tonsa and Tisbe battagliai. Tests were also conducted in the Chesapeake Bay laboratory using estuarine species including the copepod Eurytemora affinis and the inland silverside Menidia beryllina. Here, the authors report on the results of these assays, together with 1996 in situ toxicity data resulting from Norwegian field trials in the northern North Sea

  17. Split Beta-Lactamase Complementation Assay

    Indian Academy of Sciences (India)

    IAS Admin

    A Search for the Molecular Better Half! Vaishali Verma ... These assays comprise of a protein molecule, ... ciferase, beta-galactosidase, GFP, g3p of M13 filamentous ph- .... sensors of protein–protein interactions, Nature Biotechnology, Vol.20,.

  18. Linearization of the bradford protein assay.

    Science.gov (United States)

    Ernst, Orna; Zor, Tsaffrir

    2010-04-12

    Determination of microgram quantities of protein in the Bradford Coomassie brilliant blue assay is accomplished by measurement of absorbance at 590 nm. This most common assay enables rapid and simple protein quantification in cell lysates, cellular fractions, or recombinant protein samples, for the purpose of normalization of biochemical measurements. However, an intrinsic nonlinearity compromises the sensitivity and accuracy of this method. It is shown that under standard assay conditions, the ratio of the absorbance measurements at 590 nm and 450 nm is strictly linear with protein concentration. This simple procedure increases the accuracy and improves the sensitivity of the assay about 10-fold, permitting quantification down to 50 ng of bovine serum albumin. Furthermore, the interference commonly introduced by detergents that are used to create the cell lysates is greatly reduced by the new protocol. A linear equation developed on the basis of mass action and Beer's law perfectly fits the experimental data.

  19. 21 CFR 866.3210 - Endotoxin assay.

    Science.gov (United States)

    2010-04-01

    ... DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3210 Endotoxin assay. (a... intended for use in conjunction with other laboratory findings and clinical assessment of the patient to...

  20. Predicting hydrocarbon release from soil

    International Nuclear Information System (INIS)

    Poppendieck, D.; Loehr, R.C.

    2002-01-01

    'Full text:' The remediation of hazardous chemicals from soils can be a lengthy and costly process. As a result, recent regulatory initiatives have focused on risk-based corrective action (RBCA) approaches. Such approaches attempt to identify the amount of chemical that can be left at a site with contaminated soil and still be protective of human health and the environment. For hydrocarbons in soils to pose risk to human heath and the environment, the hydrocarbons must be released from the soil and accessible to microorganisms, earthworms, or other higher level organisms. The sorption of hydrocarbons to soil can reduce the availability of the hydrocarbon to receptors. Typically in soils and sediments, there is an initial fast release of a hydrocarbon from the soil to the aqueous phase followed by a slower release of the remaining hydrocarbon to the aqueous phase. The rate and extent of slow release can influence aqueous hydrocarbon concentrations and the fate and transport of hydrocarbons in the subsurface. Once the fast fraction of the chemical has been removed from the soil, the remaining fraction of a chemical may desorb at a rate that natural mechanisms can attenuate the released hydrocarbon. Hence, active remediation may be needed only until the fast fraction has been removed. However, the fast fraction is a soil and chemical specific parameter. This presentation will present a tier I type protocol that has been developed to quickly estimate the fraction of hydrocarbons that are readily released from the soil matrix to the aqueous phase. Previous research in our laboratory and elsewhere has used long-term desorption (four months) studies to determine the readily released fraction. This research shows that a single short-term (less than two weeks) batch extraction procedure provides a good estimate of the fast released fraction derived from long-term experiments. This procedure can be used as a tool to rapidly evaluate the release and bioavailability of

  1. Passive nondestructive assay of nuclear materials

    International Nuclear Information System (INIS)

    Reilly, D.; Ensslin, N.; Smith, H. Jr.; Kreiner, S.

    1991-03-01

    The term nondestructive assay (NDA) is applied to a series of measurement techniques for nuclear fuel materials. The techniques measure radiation induced or emitted spontaneously from the nuclear material; the measurements are nondestructive in that they do not alter the physical or chemical state of the nuclear material. NDA techniques are characterized as passive or active depending on whether they measure radiation from the spontaneous decay of the nuclear material or radiation induced by an external source. This book emphasizes passive NDA techniques, although certain active techniques like gamma-ray absorption densitometry and x-ray fluorescence are discussed here because of their intimate relation to passive assay techniques. The principal NDA techniques are classified as gamma-ray assay, neutron assay, and calorimetry. Gamma-ray assay techniques are treated in Chapters 1--10. Neutron assay techniques are the subject of Chapters 11--17. Chapters 11--13 cover the origin of neutrons, neutron interactions, and neutron detectors. Chapters 14--17 cover the theory and applications of total and coincidence neutron counting. Chapter 18 deals with the assay of irradiated nuclear fuel, which uses both gamma-ray and neutron assay techniques. Chapter 19 covers perimeter monitoring, which uses gamma-ray and neutron detectors of high sensitivity to check that no unauthorized nuclear material crosses a facility boundary. The subject of Chapter 20 is attribute and semiquantitative measurements. The goal of these measurements is a rapid verification of the contents of nuclear material containers to assist physical inventory verifications. Waste and holdup measurements are also treated in this chapter. Chapters 21 and 22 cover calorimetry theory and application, and Chapter 23 is a brief application guide to illustrate which techniques can be used to solve certain measurement problems

  2. Optical assay for biotechnology and clinical diagnosis.

    Science.gov (United States)

    Moczko, Ewa; Cauchi, Michael; Turner, Claire; Meglinski, Igor; Piletsky, Sergey

    2011-08-01

    In this paper, we present an optical diagnostic assay consisting of a mixture of environmental-sensitive fluorescent dyes combined with multivariate data analysis for quantitative and qualitative examination of biological and clinical samples. The performance of the assay is based on the analysis of spectrum of the selected fluorescent dyes with the operational principle similar to electronic nose and electronic tongue systems. This approach has been successfully applied for monitoring of growing cell cultures and identification of gastrointestinal diseases in humans.

  3. Calibration method for a radwaste assay system

    International Nuclear Information System (INIS)

    Dulama, C.; Dobrin, R.; Toma, Al.; Paunoiu, C.

    2004-01-01

    A waste assay system entirely designed and manufactured in the Institute for Nuclear Research is used in radwaste treatment and conditioning stream to ensure compliance with national repository radiological requirements. Usually, waste assay systems are calibrated by using various experimental arrangements including calibration phantoms. The paper presents a comparative study concerning the efficiency calibration performed by shell source method and a semiempirical, computational method based on a Monte Carlo algorithm. (authors)

  4. GEWEX SRB Shortwave Release 4

    Science.gov (United States)

    Cox, S. J.; Stackhouse, P. W., Jr.; Mikovitz, J. C.; Zhang, T.

    2017-12-01

    The NASA/GEWEX Surface Radiation Budget (SRB) project produces shortwave and longwave surface and top of atmosphere radiative fluxes for the 1983-near present time period. Spatial resolution is 1 degree. The new Release 4 uses the newly processed ISCCP HXS product as its primary input for cloud and radiance data. The ninefold increase in pixel number compared to the previous ISCCP DX allows finer gradations in cloud fraction in each grid box. It will also allow higher spatial resolutions (0.5 degree) in future releases. In addition to the input data improvements, several important algorithm improvements have been made since Release 3. These include recalculated atmospheric transmissivities and reflectivities yielding a less transmissive atmosphere. The calculations also include variable aerosol composition, allowing for the use of a detailed aerosol history from the Max Planck Institut Aerosol Climatology (MAC). Ocean albedo and snow/ice albedo are also improved from Release 3. Total solar irradiance is now variable, averaging 1361 Wm-2. Water vapor is taken from ISCCP's nnHIRS product. Results from GSW Release 4 are presented and analyzed. Early comparison to surface measurements show improved agreement.

  5. Aluminum corrosion product release kinetics

    Energy Technology Data Exchange (ETDEWEB)

    Edwards, Matt, E-mail: Matthew.Edwards@cnl.ca; Semmler, Jaleh; Guzonas, Dave; Chen, Hui Qun; Toor, Arshad; Hoendermis, Seanna

    2015-07-15

    Highlights: • Release of Al corrosion product was measured in simulated post-LOCA sump solutions. • Increased boron was found to enhance Al release kinetics at similar pH. • Models of Al release as functions of time, temperature, and pH were developed. - Abstract: The kinetics of aluminum corrosion product release was examined in solutions representative of post-LOCA sump water for both pressurized water and pressurized heavy-water reactors. Coupons of AA 6061 T6 were exposed to solutions in the pH 7–11 range at 40, 60, 90 and 130 °C. Solution samples were analyzed by inductively coupled plasma atomic emission spectroscopy, and coupon samples were analyzed by secondary ion mass spectrometry. The results show a distinct “boron effect” on the release kinetics, expected to be caused by an increase in the solubility of the aluminum corrosion products. New models were developed to describe both sets of data as functions of temperature, time, and pH (where applicable)

  6. Controlled Release Formulations of Auxinic Herbicides

    Science.gov (United States)

    Kowalski, Witold J.; Siłowiecki, Andrzej.; Romanowska, Iwona; Glazek, Mariola; Bajor, Justyna; Cieciwa, Katarzyna; Rychter, Piotr

    2013-04-01

    Controlled release formulations are applied extensively for the release of active ingredients such as plant protection agents and fertilizers in response to growing concern for ecological problems associated with increased use of plant protection chemicals required for intensive agricultural practices [1]. We synthesized oligomeric mixtures of (R,S)-3-hydroxy butyric acid chemically bonded with 2,4-D, Dicamba and MCPA herbicides (HBA) respectively, and determined their molecular structure and molecular weight dispersion by the size exclusion chromatography, proton magnetic resonance spectrometry and electro-spray ionization mass spectrometry. Further we carried out bioassays of herbicidal effectiveness of the HBA herbicides vs. series of dicotyledonous weeds and crop injury tests [2, 3, 4]. Field bioassays were accomplished according to the EPPO standards [5]. Groups of representative weeds (the development stages in the BCCH scale: 10 - 30) were selected as targets. Statistical variabilities were assessed by the Fisher LSD test for plants treated with the studied herbicides in form of HBA oligomers, the reference herbicides in form of dimethyl ammonium salts (DMA), and untreated plants. No statistically significant differences in the crop injuries caused by the HBA vs. the DMA reference formulation were observed. The effectiveness of the HBA herbicides was lower through the initial period (ca. 2 weeks) relative to the DMA salts, but a significant increase in the effectiveness of the HBA systems followed during the remaining fraction of each assay. After 6 weeks all observed efficiencies approached 100%. The death of weeds treated with the HBA herbicides was delayed when compared with the DMA reference herbicides. The delayed uptake observed for the HBA oligomers relative to the DMA salts was due to controlled release phenomena. In case of the DMA salts the total amount of active ingredients was available at the target site. By contrast, the amount of an active

  7. Nondestructive assay methods for irradiated nuclear fuels

    International Nuclear Information System (INIS)

    Hsue, S.T.; Crane, T.W.; Talbert, W.L. Jr.; Lee, J.C.

    1978-01-01

    This report is a review of the status of nondestructive assay (NDA) methods used to determine burnup and fissile content of irradiated nuclear fuels. The gamma-spectroscopy method measures gamma activities of certain fission products that are proportional to the burnup. Problems associated with this method are migration of the fission products and gamma-ray attenuation through the relatively dense fuel material. The attenuation correction is complicated by generally unknown activity distributions within the assemblies. The neutron methods, which usually involve active interrogation and prompt or delayed signal counting, are designed to assay the fissile content of the spent-fuel elements. Systems to assay highly enriched spent-fuel assemblies have been tested extensively. Feasibility studies have been reported of systems to assay light-water reactor spent-fuel assemblies. The slowing-down spectrometer and neutron resonance absorption methods can distinguish between the uranium and plutonium fissile contents, but they are limited to the assay of individual rods. We have summarized the status of NDA techniques for spent-fuel assay and present some subjects in need of further investigation. Accuracy of the burnup calculations for power reactors is also reviewed

  8. Development of a Rapid Insulin Assay by Homogenous Time-Resolved Fluorescence.

    Directory of Open Access Journals (Sweden)

    Zachary J Farino

    Full Text Available Direct measurement of insulin is critical for basic and clinical studies of insulin secretion. However, current methods are expensive and time-consuming. We developed an insulin assay based on homogenous time-resolved fluorescence that is significantly more rapid and cost-effective than current commonly used approaches. This assay was applied effectively to an insulin secreting cell line, INS-1E cells, as well as pancreatic islets, allowing us to validate the assay by elucidating mechanisms by which dopamine regulates insulin release. We found that dopamine functioned as a significant negative modulator of glucose-stimulated insulin secretion. Further, we showed that bromocriptine, a known dopamine D2/D3 receptor agonist and newly approved drug used for treatment of type II diabetes mellitus, also decreased glucose-stimulated insulin secretion in islets to levels comparable to those caused by dopamine treatment.

  9. Digital Drug Dosing: Dosing in Drug Assays by Light-Defined Volumes of Hydrogels with Embedded Drug-Loaded Nanoparticles

    DEFF Research Database (Denmark)

    Faralli, Adele; Melander, Fredrik; Larsen, Esben Kjær Unmack

    2014-01-01

    Polyethylene glycol (PEG)-based hydrogels are widely used for biomedical applications, including matrices for controlled drug release. We present a method for defining drug dosing in screening assays by light-activated cross-linking of PEG-diacrylate hydrogels with embedded drug-loaded liposome...

  10. Training Materials for Release 3

    DEFF Research Database (Denmark)

    Wake, Jo Dugstad; Hansen, Cecilie; Debus, Kolja

    This document, D7.4 – training materials for release 3, provides an overview of the training material for version 3 of the NEXT-TELL tools and methods. Previous documents submitted as part of work package 7, which is about teacher training, are D7.1 – Training Concept, D7.2 – Training Materials...... for Release 1 and D7.3 – Training Materials for Release 2. D7.4 builds on D7.1 and D7.2 and D7.3. D7.4 contains further development of previous work within WP7, essentially a revised theoretical approach to the teacher training, and expansion of the notion of tool training. The media in use have been expanded...

  11. Controlled Release from Recombinant Polymers

    Science.gov (United States)

    Price, Robert; Poursaid, Azadeh; Ghandehari, Hamidreza

    2014-01-01

    Recombinant polymers provide a high degree of molecular definition for correlating structure with function in controlled release. The wide array of amino acids available as building blocks for these materials lend many advantages including biorecognition, biodegradability, potential biocompatibility, and control over mechanical properties among other attributes. Genetic engineering and DNA manipulation techniques enable the optimization of structure for precise control over spatial and temporal release. Unlike the majority of chemical synthetic strategies used, recombinant DNA technology has allowed for the production of monodisperse polymers with specifically defined sequences. Several classes of recombinant polymers have been used for controlled drug delivery. These include, but are not limited to, elastin-like, silk-like, and silk-elastinlike proteins, as well as emerging cationic polymers for gene delivery. In this article, progress and prospects of recombinant polymers used in controlled release will be reviewed. PMID:24956486

  12. Release of Growth Factors into Root Canal by Irrigations in Regenerative Endodontics.

    Science.gov (United States)

    Zeng, Qian; Nguyen, Sean; Zhang, Hongming; Chebrolu, Hari Priya; Alzebdeh, Dalia; Badi, Mustafa A; Kim, Jong Ryul; Ling, Junqi; Yang, Maobin

    2016-12-01

    The aim of this study was to investigate the release of growth factors into root canal space after the irrigation procedure of regenerative endodontic procedure. Sixty standardized root segments were prepared from extracted single-root teeth. Nail varnish was applied to all surfaces except the root canal surface. Root segments were irrigated with 1.5% NaOCl + 17% EDTA, 2.5% NaOCl + 17% EDTA, 17% EDTA, or deionized water. The profile of growth factors that were released after irrigation was studied by growth factor array. Enzyme-linked immunosorbent assay was used to validate the release of transforming growth factor (TGF)-β1 and basic fibroblast growth factor (bFGF) at 4 hours, 1 day, and 3 days after irrigation. The final concentrations were calculated on the basis of the root canal volume measured by cone-beam computed tomography. Dental pulp stem cell migration on growth factors released from root segments was measured by using Transwell assay. Total of 11 of 41 growth factors were detected by growth factors array. Enzyme-linked immunosorbent assay showed that TGF-β1 was released in all irrigation groups. Compared with the group with 17% EDTA (6.92 ± 4.49 ng/mL), the groups with 1.5% NaOCl + 17% EDTA and 2.5% NaOCl + 17% EDTA had significantly higher release of TGF-β1 (69.04 ± 30.41 ng/mL and 59.26 ± 3.37 ng/mL, respectively), with a peak release at day 1. The release of bFGF was detected at a low level in all groups (0 ng/mL to 0.43 ± 0.22 ng/mL). Migration assay showed the growth factors released from root segments induced dental pulp stem cell migration. The root segment model in present study simulated clinical scenario and indicated that the current irrigation protocol released a significant amount of TGF-β1 but not bFGF. The growth factors released into root canal space induced dental pulp stem cell migration. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  13. Solid-phase peptide quantitation assay using labeled monoclonal antibody and glutaraldehyde fixation

    International Nuclear Information System (INIS)

    Kasprzyk, P.G.; Cuttitta, F.; Avis, I.; Nakanishi, Y.; Treston, A.; Wong, H.; Walsh, J.H.; Mulshine, J.L.

    1988-01-01

    A solid-phase radioimmunoassay utilizing iodinated peptide-specific monoclonal antibody as a detection system instead of labeled peptide has been developed. Regional specific monoclonal antibodies to either gastrin-releasing peptide or gastrin were used as models to validate the general application of our modified assay. Conditions for radioactive labeling of the monoclonal antibody were determined to minimize oxidant damage, which compromises the sensitivity of other reported peptide quantitation assays. Pretreatment of 96-well polyvinyl chloride test plates with a 5% glutaraldehyde solution resulted in consistent retention of sufficient target peptide on the solid-phase matrix to allow precise quantitation. This quantitative method is completed within 1 h of peptide solid phasing. Pretreatment of assay plates with glutaraldehyde increased binding of target peptide and maximized antibody binding by optimizing antigen presentation. The hypothesis that glutaraldehyde affects both peptide binding to the plate and orientation of the peptide was confirmed by analysis of several peptide analogs. These studies indicate that peptide binding was mediated through a free amino group leaving the carboxy-terminal portion of the target peptide accessible for antibody binding. It was observed that the length of the peptide also affects the amount of monoclonal antibody that will bind. Under the optimal conditions, results from quantitation of gastrin-releasing peptide in relevant samples agree well with those from previously reported techniques. Thus, we report here a modified microplate assay which may be generally applied for the rapid and sensitive quantitation of peptide hormones

  14. Use of the local lymph node assay in assessment of immune function

    International Nuclear Information System (INIS)

    Berg, Femke A. van den; Baken, Kirsten A.; Vermeulen, Jolanda P.; Gremmer, Eric R.; Steeg, Harry van; Loveren, Henk van

    2005-01-01

    The murine local lymph node assay (LLNA) was originally developed as a predictive test method for the identification of chemicals with sensitizing potential. In this study we demonstrated that an adapted LLNA can also be used as an immune function assay by studying the effects of orally administered immunomodulating compounds on the T-cell-dependent immune response induced by the contact sensitizer 2,4-dinitrochlorobenzene (DNCB). C57Bl/6 mice were treated with the immunotoxic compounds cyclosporin A (CsA), bis(tri-n-butyltin)oxide (TBTO) or benzo[a]pyrene (B[a]P). Subsequently, cell proliferation and interferon-γ (IFN-γ) and interleukin (IL)-4 release were determined in the auricular lymph nodes (LNs) after DNCB application on both ears. Immunosuppression induced by CsA, TBTO and B[a]P was clearly detectable in this application of the LLNA. Cytokine release measurements proved valuable to confirm the results of the cell proliferation assay and to obtain an indication of the effect on Th1/Th2 balance. We believe to have demonstrated the applicability of an adapted LLNA as an immune function assay in the mouse

  15. Nanostructured Diclofenac Sodium Releasing Material

    Science.gov (United States)

    Nikkola, L.; Vapalahti, K.; Harlin, A.; Seppälä, J.; Ashammakhi, N.

    2008-02-01

    Various techniques have been developed to produce second generation biomaterials for tissue repair. These include extrusion, molding, salt leaching, spinning etc, but success in regenerating tissues has been limited. It is important to develop porous material, yet with a fibrous structure for it to be biomimetic. To mimic biological tissues, the extra-cellular matrix usually contains fibers in nano scale. To produce nanostructures, self-assembly or electrospinning can be used. Adding a drug release function to such a material may advance applications further for use in controlled tissue repair. This turns the resulting device into a multifunctional porous, fibrous structure to support cells and drug releasing properties in order to control tissue reactions. A bioabsorbable poly(ɛ-caprolactone-co-D,L lactide) 95/5 (PCL) was made into diluted solution using a solvent, to which was added 2w-% of diclofenac sodium (DS). Nano-fibers were made by electrospinning onto substrate. Microstructure of the resulting nanomat was studied using SEM and drug release profiles with UV/VIS spectroscopy. Thickness of the electrospun nanomat was about 2 mm. SEM analysis showed that polymeric nano-fibers containing drug particles form a highly interconnected porous nano structure. Average diameter of the nano-fibers was 130 nm. There was a high burst peak in drug release, which decreased to low levels after one day. The used polymer has slow a degradation rate and though the nanomat was highly porous with a large surface area, drug release rate is slow. It is feasible to develop a nano-fibrous porous structure of bioabsorbable polymer, which is loaded with test drug. Drug release is targeted at improving the properties of biomaterial for use in controlled tissue repair and regeneration.

  16. Blood histamine release: A new allergy blood test

    International Nuclear Information System (INIS)

    Faraj, B.A.; Gottlieb, G.R.; Camp, V.M.; Lollies, P.

    1985-01-01

    Allergen-mediated histamine release from human leukocytes represents an important model for in vitro studies of allergic reactions. The purpose of this study was to determine whether the measurement of histamine released in allergic patients (pts) by radioenzymatic assay following mixing of their blood with common allergens represents a reliable index for diagnosis of atopic allergy. Three categories of allergies were used: (1) housedust and mite; (2) cat and dog dander; (3) trees and grasses and ragweed mixture. The presence of allergy was established by intradermal skin testing in the study group of 82 pts. Significant atopy was defined as ≥ 3+ (overall range 0-4 +, negative to maximum) on skin testing. The test was carried out in tubes with 0.5 ml heparinized blood, 0.5 ml tris albumin buffer, and one of the allergens (60-100 PNU/ml). In 20 controls without allergy, there always was ≤ 4% histamine release (normal response). A significant allergen-mediated histamine release, ranging from 12 to 30% of the total blood histamine content, was observed in 96% of the pts with skin test sensitivity of ≥ 3+. There was good agreement between skin testing and histamine release in terms of the allergen causing the response. Thus, measurement of histamine release in blood in response to allergen challenge represents a clinically useful in vitro test for the diagnosis of atopic allergy. Because data can be obtained from a single sample and are highly quantitative, this new method should have application to the longitudinal study of allergic pts and to the assessment of interventions

  17. Limited Releases of Krsko NPP

    International Nuclear Information System (INIS)

    Breznik, B.; Kovac, A.

    2001-01-01

    Full text: Krsko Nuclear Power Plant is about 700 MW Pressurised Water Reactor plant located in Slovenia close to the border with Croatia. The authorised limit for the radioactive releases is basically set to 50 μSv effective dose per year to the members of the public. There is also additional limitation of total activities released in a year and concentration. The poster presents the effluents of the year 2000 and evaluated dose referring to the limits and to the natural and other sources of radiation around the plant. (author)

  18. Formulation of Dipyridamole Sustained Release Tablet Using Floating System

    Directory of Open Access Journals (Sweden)

    Lenny Mauilida Valentina

    2011-06-01

    Full Text Available Dipyridamole is a drug for prevention of postoperative thromboembolic complication of heart valve replacement and long term therapy of angina pectoris will be well absorbed in stomach. To maintain therapeutic plasma concentration in long time and to increase bioavalaibility is needed a sustained release dosage form having the long residence time in the stomach. The objective of this research was to make floating sustained release tablet of dipyridamole conforming to the requirement that was set up by dipyridamol therapeutic concentration. Tablets were made by wet granulation method using aquadest as a liquid binder, HPMC K4M, Ac-di-sol, Avicel PH 102, talk, and Mg stearat. Dissolution assay was carried out using type 2 release tester at rotation speed of 50 rpm in medium 900 mL HCl 0.1 N at 37 ± 0.5 °C for 8 hours. The formulation containing of 50 mg dipirydamole, HPMC K4M (30%, Ac-di-sol (20%, Avicel PH 102 (37%, talk (2%, and Mg stearat (1% released 59.61 ± 6.73% and 89.34 ± 5.87% of dipyridamole respectively after 4 and 8 hours that conformed to the requirement.

  19. Microbubbles-Assisted Ultrasound Triggers the Release of Extracellular Vesicles

    Directory of Open Access Journals (Sweden)

    Yuana Yuana

    2017-07-01

    Full Text Available Microbubbles-assisted ultrasound (USMB has shown promise in improving local drug delivery. The formation of transient membrane pores and endocytosis are reported to be enhanced by USMB, and they contribute to cellular drug uptake. Exocytosis also seems to be linked to endocytosis upon USMB treatment. Based on this rationale, we investigated whether USMB triggers exocytosis resulting in the release of extracellular vesicles (EVs. USMB was performed on a monolayer of head-and-neck cancer cells (FaDu with clinically approved microbubbles and commonly used ultrasound parameters. At 2, 4, and 24 h, cells and EV-containing conditioned media from USMB and control conditions (untreated cells, cells treated with microbubbles and ultrasound only were harvested. EVs were measured using flow cytometric immuno-magnetic bead capture assay, immunogold electron microscopy, and western blotting. After USMB, levels of CD9 exposing-EVs significantly increased at 2 and 4 h, whereas levels of CD63 exposing-EVs increased at 2 h. At 24 h, EV levels were comparable to control levels. EVs released after USMB displayed a heterogeneous size distribution profile (30–1200 nm. Typical EV markers CD9, CD63, and alix were enriched in EVs released from USMB-treated FaDu cells. In conclusion, USMB treatment triggers exocytosis leading to the release of EVs from FaDu cells.

  20. Bubble Jet agent release cartridge for chemical single cell stimulation.

    Science.gov (United States)

    Wangler, N; Welsche, M; Blazek, M; Blessing, M; Vervliet-Scheebaum, M; Reski, R; Müller, C; Reinecke, H; Steigert, J; Roth, G; Zengerle, R; Paust, N

    2013-02-01

    We present a new method for the distinct specific chemical stimulation of single cells and small cell clusters within their natural environment. By single-drop release of chemical agents with droplets in size of typical cell diameters (d agent release cartridge with integrated fluidic structures and integrated agent reservoirs are shown, tested, and compared in this publication. The single channel setup features a fluidic structure fabricated by anisotropic etching of silicon. To allow for simultaneous release of different agents even though maintaining the same device size, the second type comprises a double channel fluidic structure, fabricated by photolithographic patterning of TMMF. Dispensed droplet volumes are V = 15 pl and V = 10 pl for the silicon and the TMMF based setups, respectively. Utilizing the agent release cartridges, the application in biological assays was demonstrated by hormone-stimulated premature bud formation in Physcomitrella patens and the individual staining of one single L 929 cell within a confluent grown cell culture.

  1. Selection of non-destructive assay methods: Neutron counting or calorimetric assay?

    International Nuclear Information System (INIS)

    Cremers, T.L.; Wachter, J.R.

    1994-01-01

    The transition of DOE facilities from production to D ampersand D has lead to more measurements of product, waste, scrap, and other less attractive materials. Some of these materials are difficult to analyze by either neutron counting or calorimetric assay. To determine the most efficacious analysis method, variety of materials, impure salts and hydrofluorination residues have been assayed by both calorimetric assay and neutron counting. New data will be presented together with a review of published data. The precision and accuracy of these measurements are compared to chemistry values and are reported. The contribution of the gamma ray isotopic determination measurement to the overall error of the calorimetric assay or neutron assay is examined and discussed. Other factors affecting selection of the most appropriate non-destructive assay method are listed and considered

  2. Controlling variation in the comet assay

    Directory of Open Access Journals (Sweden)

    Andrew Richard Collins

    2014-10-01

    Full Text Available Variability of the comet assay is a serious issue, whether it occurs from experiment to experiment in the same laboratory, or between different laboratories analysing identical samples. Do we have to live with high variability, just because the comet assay is a biological assay rather than analytical chemistry? Numerous attempts have been made to limit variability by standardising the assay protocol, and the critical steps in the assay have been identified; agarose concentration, duration of alkaline incubation, and electrophoresis conditions (time, temperature and voltage gradient are particularly important. Even when these are controlled, variation seems to be inevitable. It is helpful to include in experiments reference standards, i.e. cells with a known amount of specific damage to the DNA. They can be aliquots frozen from a single large batch of cells, either untreated (negative controls or treated with, for example, H2O2 or X-rays to induce strand breaks (positive control for the basic assay, or photosensitiser plus light to oxidise guanine (positive control for Fpg- or OGG1-sensitive sites. Reference standards are especially valuable when performing a series of experiments over a long period - for example, analysing samples of white blood cells from a large human biomonitoring trial - to check that the assay is performing consistently, and to identify anomalous results necessitating a repeat experiment. The reference values of tail intensity can also be used to iron out small variations occurring from day to day. We present examples of the use of reference standards in human trials, both within one laboratory and between different laboratories, and describe procedures that can be used to control variation.

  3. Cell Culture Assay for Human Noroviruses [response

    Energy Technology Data Exchange (ETDEWEB)

    Straub, Tim M.; Honer Zu Bentrup, Kerstin; Orosz Coghlan, Patricia; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza A.; Nickerson, Cheryl A.

    2007-07-01

    We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools

  4. The Comet Assay: Tails of the (Unexpected. Use of the comet assay in pharmaceutical development.

    Directory of Open Access Journals (Sweden)

    Bas-jan Van Der Leede

    2015-08-01

    Full Text Available In genotoxicity testing of pharmaceuticals the rodent alkaline comet assay is being increasingly used as a second in vivo assay in addition to the in vivo micronucleus assay to mitigate in vitro positive results as recommended by regulatory guidance. In this presentation we want to give insight into the circumstances in vivo comet assay is deployed in a Genetic Toxicology Department of a pharmaceutical company. As the in vivo comet assay is a salvage assay, it means that some events have occurred in an in vitro assay and that the compound (or metabolite responsible for this signal is potentially deselected for further development. More than often the decision to perform an in vivo comet assay is at a very early stage in development and the first time that the compound will be tested in vivo at high/toxic dose levels. As almost no toxicokinetic data and tissue distribution data are available a careful design with maximizes the chances for successful mitigation is necessary. Decisions on acute or repeated dosing need to be made and arrangements for combining the in vivo comet assay with the in vivo micronucleus assay are to be considered. Often synthesis methods need to be scaled up fast to provide the required amount of compound and information on suitable formulations needs to be in place. As exposure data is crucial for interpretation of results, analytical methods need to be brought in place rapidly. An experienced multi skilled and communicative team needs to be available to deploy successfully this kind of assays at an early stage of development. We will present a few scenarios on study conduct and demonstrate how this assay can make a difference for the further development of a new drug.

  5. Random assay in radioimmunoassay: Feasibility and application compared with batch assay

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jung Min; Lee, Hwan Hee; Park, Sohyun; Kim, Tae Sung; Kim, Seok Ki [Dept. of Nuclear MedicineNational Cancer Center, Goyang (Korea, Republic of)

    2016-12-15

    The batch assay has been conventionally used for radioimmunoassay (RIA) because of its technical robustness and practical convenience. However, it has limitations in terms of the relative lag of report time due to the necessity of multiple assays in a small number of samples compared with the random assay technique. In this study, we aimed to verify whether the random assay technique can be applied in RIA and is feasible in daily practice. The coefficients of variation (CVs) of eight standard curves within a single kit were calculated in a CA-125 immunoradiometric assay (IRMA) for the reference of the practically ideal CV of the CA-125 kit. Ten standard curves of 10 kits from 2 prospectively collected lots (pLot) and 85 standard curves of 85 kits from 3 retrospectively collected lots (Lot) were obtained. Additionally, the raw measurement data of both 170 control references and 1123 patients' sera were collected retrospectively between December 2015 and January 2016. A standard curve of the first kit of each lot was used as a master standard curve for a random assay. The CVs of inter-kits were analyzed in each lot, respectively. All raw measurements were normalized by decay and radioactivity. The CA-125 values from control samples and patients' sera were compared using the original batch assay and random assay. In standard curve analysis, the CVs of inter-kits in pLots and Lots were comparable to those within a single kit. The CVs from the random assay with normalization were similar to those from the batch assay in the control samples (CVs % of low/high concentration; Lot1 2.71/1.91, Lot2 2.35/1.83, Lot3 2.83/2.08 vs. Lot1 2.05/1.21, Lot2 1.66/1.48, Lot3 2.41/2.14). The ICCs between the batch assay and random assay using patients' sera were satisfactory (Lot1 1.00, Lot2 0.999, Lot3 1.00). The random assay technique could be successfully applied to the conventional CA-125 IRMA kits. The random assay showed strong agreement with the batch assay. The

  6. Energy Release in Solar Flares,

    Science.gov (United States)

    1982-10-01

    Plasma Research, Stanford University P. Kaufmanu CRAA/CNPq -Conseiho lacional de Desenvolvimento Cientifico e Tecnologico, Slo Paulo, SP, Brasil D.F...three phases of energy release in solar flares (Sturrock, 1980). However, a recent article by Feldman e a.. (1982) points to a significant

  7. Lignin based controlled release coatings

    NARCIS (Netherlands)

    Mulder, W.J.; Gosselink, R.J.A.; Vingerhoeds, M.H.; Harmsen, P.F.H.; Eastham, D.

    2011-01-01

    Urea is a commonly used fertilizer. Due to its high water-solubility, misuse easily leads to excess nitrogen levels in the soil. The aim of this research was to develop an economically feasible and biodegradable slow-release coating for urea. For this purpose, lignin was selected as coating

  8. Controlled Release from Zein Matrices

    NARCIS (Netherlands)

    Bouman, Jacob; Belton, Peter; Venema, Paul; Linden, Van Der Erik; Vries, De Renko; Qi, Sheng

    2016-01-01

    Purpose: In earlier studies, the corn protein zein is found to be suitable as a sustained release agent, yet the range of drugs for which zein has been studied remains small. Here, zein is used as a sole excipient for drugs differing in hydrophobicity and isoelectric point: indomethacin,

  9. Dry release of suspended nanostructures

    DEFF Research Database (Denmark)

    Forsén, Esko Sebastian; Davis, Zachary James; Dong, M.

    2004-01-01

    , the technique enables long time storage and transportation of produced devices without the risk of stiction. By combining the dry release method with a plasma deposited anti-stiction coating both fabrication induced stiction, which is mainly caused by capillary forces originating from the dehydration...

  10. Comet assay on mice testicular cells

    Directory of Open Access Journals (Sweden)

    Anoop Kumar Sharma

    2015-05-01

    Full Text Available Heritable mutations may result in a variety of adverse outcomes including genetic disease in the offspring. In recent years the focus on germ cell mutagenicity has increased and the “Globally Harmonized System of Classification and Labelling of Chemicals (GHS” has published classification criteria for germ cell mutagens (Speit et al., 2009. The in vivo Comet assay is considered a useful tool for investigating germ cell genotoxicity. In the present study DNA strand breaks in testicular cells of mice were investigated. Different classes of chemicals were tested in order to evaluate the sensitivity of the comet assay in testicular cells. The chemicals included environmentally relevant substances such as Bisphenol A, PFOS and Tetrabrombisphenol A. Statistical power calculations will be presented to aid in the design of future Comet assay studies on testicular cells. Power curves were provided with different fold changes in % tail DNA, different number of cells scored and different number of gels (Hansen et al., 2014. An example is shown in Figure 1. A high throughput version of the Comet assay was used. Samples were scored with a fully automatic comet assay scoring system that provided faster scoring of randomly selected cells.

  11. Immune chromatography: a quantitative radioimmunological assay

    International Nuclear Information System (INIS)

    Davis, J.W.; Demetriades, M.; Bowen, J.M.

    1984-01-01

    Immune chromatography, a radioimmunological binding assay, employs paper chromatography to separate immune complexes from free antigen and antibodies. During chromatography free antigen and antibodies become distributed throughout the paper, while immune complexes remain near the bottoms of the strips. The chromatographic differences can be made quantitative by using either iodinated antigens or antibodies. Under these conditions nanogram quantities of antigen can be detected or antibodies in sera diluted several 1000-fold. The immune chromatography assay can also be performed as an indirect assay, since the paper strips are cut from nitrocellulose paper. In this case the immune components are absorbed by the paper during chromatography. Antigen is then detected with an iodinated second antibody. The indirect immune chromatography assay is particularly useful for identifying different sera that react with the same antigen. Reaction with the first serum before chromatography reduces the amount of antigen available to the second serum following chromatography. In addition to characterizing the immune chromatography procedure, we discuss the possible applications of chromatography assays for the quantitation of other types of molecular binding interactions. (Auth.)

  12. Evaluation of three gentamicin serum assay techniques

    International Nuclear Information System (INIS)

    Matzke, G.R.; Gwizdala, C.; Wery, J.; Ferry, D.; Starnes, R.

    1982-01-01

    This investigation was designed to compare the enzyme-modified immunoassay (Syva--EMIT) with a radioimmunoassay (New England Nuclear--RIA) and the radiometric assay (Johnston--BACTEC) to determine the optimal assay for use in our aminoglycoside dosing service. The serum concentration determinations obtained via the three assay methods were analyzed by linear regression analysis. Significant positive correlations were noted between the three assay techniques (p less than 0.005) during both sample collection phases. The coefficients of determination for EMIT vs BACTEC and RIA vs BACTEC were 0.73 and 0.83 during phase 1, respectively, and 0.65 and 0.68 during phase 2, respectively. The slope of the regression lines also varied markedly during the two phases; 0.49 and 0.42 for EMIT and for RIA vs BACTEC, respectively, during phase 1 compound with 1.12 and 0.77, respectively, during phase 2. The differences noted in these relationships during phase 1 and 2 may be related to the alteration of the pH of the control sera utilized in the BACTEC assay. In contrast, RIA vs EMIT regression analysis indicated that existence of a highly significant relationship (p less than 0.0005 and r2 . 0.90). The EMIT technique was the easiest and most accurate for determination of serum gentamicin concentrations, whereas the BACTEC method was judged unacceptable for clinical use

  13. Pilot study of human recombinant interferon gamma and accelerated hyperfractionated thoracic radiation therapy in patients with unresectable stage IIIA/B nonsmall cell lung cancer

    International Nuclear Information System (INIS)

    Shaw, Edward G.; Deming, Richard L.; Creagan, Edward T.; Nair, Suresh; Su, John Q.; Levitt, Ralph; Steen, Preston D.; Wiesenfeld, Martin; Mailliard, James A.

    1995-01-01

    Purpose: Gamma interferon has a wide range of properties, including the ability to sensitize solid tumor cells to the effects of ionizing radiation. The North Central Cancer Treatment Group has previously completed pilot studies of accelerated hyperfractionated thoracic radiation therapy (AHTRT) in patients with unresectable Stage IIIA/B nonsmall cell lung cancer (NSCLC). This Phase I study was designed to assess the toxicity of concomitant gamma interferon and AHTRT in a similar patient population. Methods and Materials: Between December 1991 and May 1992, 18 patients with unresectable Stage IIIA/B NSCLC were treated with daily gamma interferon (0.2 mg subcutaneously) concomitant with AHTRT (60 Gy given in 1.5 Gy twice daily fractions). All patients had an Eastern Cooperative Oncology Group performance status of 0 or 1 with weight loss < 5%. Eight patients had Stage IIIA and 10 had Stage IIIB disease. Results: Nine patients (50%) experienced severe, life-threatening, or fatal toxicities. Eight of the patients (44%) developed significant radiation pneumonitis, which was severe in six patients and fatal in two patients (11% treatment-related mortality). Two patients (11%) developed severe radiation esophagitis. With follow-up of 15-21 months, 2 patients are alive, and 16 have died. The median survival time and 1-year survival rate is 7.8 months and 38%, respectively. Conclusion: Gamma interferon appeared to sensitize normal lung tissue to the effects of radiation, as demonstrated by the high incidence of severe or fatal radiation pneumonitis. We do not recommend pursuing gamma interferon as a radiosensitizer in this setting

  14. No changes in serum concentrations of interleukin 10 (IL-10) and interferon gamma (IF-gamma) before and after treatment of the thyroid eye disease (TED).

    Science.gov (United States)

    Laban-Guceva, Nevenka; Antova, Magdalena; Bogoev, Milco

    2007-11-01

    TED is a severe eye disease leading in rare cases to decrease of sight, optic nerve compression and blindness. Recently, significant progresses in understanding the disease have been done. Nevertheless, the treatment of the disease, especially in its severe form remains challenging. Glucocorticoids (GC) have been the basis of the treatment for a long time. Orbital irradiation (OI) and optical decompression (OD) are also used in managing the severe forms of TED. Somatostatin, intravenous immunoglobulin have been also used, with conflicting results. Regarding the potential for the treatment of TED with cytokine antagonists, controlled clinical studies are not available. Since cytokines play an important role in the pathogenesis of the TED, they seemed to be logical choice for modern TED treatment. It has been shown that both Th1 (interleukin-2, tumor necrosis factor gamma, interleukin gamma) and Th2 (interleukin -4, -5, -10) profile T cells are activated in the TED. We therefore measured interleukin-gamma, IF-gamma and interleukin -10 (IL-10)(Th1 and Th2 pattern) to assess its relationship to the course of the disease. This paper shows that both Th1 (IL-2) and Th2 (IF-gamma) pathways represented by those two cytokines are not involved (IL-10 before 2.29+/-5.23 and after treatment 3.77+/-8.44; IF gamma before 0.50+/-0.24 and after treatment 0.35+/-0.19). No relationship to the response to treatment was found. GC resulted in positive response in 8/22 patients, OI (12 patients) given after CS therapy, resulted in a response in all patients. Increase in proptosis, loss of visual acuity is spite of CS treatment prompted OD in two patients, who both recovered visual acuity and proptosis fell under 25 mm Hertel.

  15. Knockdown of menin affects pre-mRNA processing and promoter fidelity at the interferon-gamma inducible IRF1 gene

    Directory of Open Access Journals (Sweden)

    Auriemma Lauren B

    2012-01-01

    Full Text Available Abstract Background The tumor suppressor menin (MEN1 is mutated in the inherited disease multiple endocrine neoplasia type I, and has several documented cellular roles, including the activation and repression of transcription effected by several transcription factors. As an activator, MEN1 is a component of the Set1-like mixed lineage leukemia (MLL MLL1/MLL2 methyltransferase complex that methylates histone H3 lysine 4 (H3K4. MEN1 is localized to the signal transducer and activator of transcription 1 (STAT1-dependent gene, interferon regulatory factor 1 (IRF1, and is further recruited when IRF1 transcription is triggered by interferon-γ signaling. Results RNAi-mediated knockdown of MEN1 alters the H3K4 dimethylation and H3 acetylation profiles, and the localization of histone deacetylase 3, at IRF1. While MEN1 knockdown does not impact the rate of transcription, IRF1 heteronuclear transcripts become enriched in MEN1-depleted cells. The processed mRNA and translated protein product are concomitantly reduced, and the antiviral state is attenuated. Additionally, the transcription start site at the IRF1 promoter is disrupted in the MEN1-depleted cells. The H3K4 demethylase, lysine specific demethylase 1, is also associated with IRF1, and its inhibition alters H3K4 methylation and disrupts the transcription start site as well. Conclusions Taken together, the data indicate that MEN1 contributes to STAT1-activated gene expression in a novel manner that includes defining the transcription start site and RNA processing.

  16. Redesigning Protein Cavities as a Strategy for Increasing Affinity in Protein-Protein Interaction: Interferon-gamma Receptor 1 as a Model

    Czech Academy of Sciences Publication Activity Database

    Černý, Jiří; Biedermannová, Lada; Mikulecký, Pavel; Zahradník, Jiří; Charnavets, Tatsiana; Šebo, Peter; Schneider, Bohdan

    2015-01-01

    Roč. 716945, 28 April 2015 (2015) ISSN 2314-6133 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0109; GA ČR GAP305/10/2184 Institutional support: RVO:86652036 Keywords : CONFORMATIONAL ENTROPY * MOLECULAR RECOGNITION * BINDING-AFFINITY Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.134, year: 2015

  17. The Rhoptry Proteins ROP18 and ROP5 Mediate Toxoplasma gondii Evasion of the Murine, But Not the Human, Interferon-Gamma Response

    Science.gov (United States)

    Niedelman, Wendy; Gold, Daniel A.; Rosowski, Emily E.; Sprokholt, Joris K.; Lim, Daniel; Farid Arenas, Ailan; Melo, Mariane B.; Spooner, Eric; Yaffe, Michael B.; Saeij, Jeroen P. J.

    2012-01-01

    The obligate intracellular parasite Toxoplasma gondii secretes effector proteins into the host cell that manipulate the immune response allowing it to establish a chronic infection. Crosses between the types I, II and III strains, which are prevalent in North America and Europe, have identified several secreted effectors that determine strain differences in mouse virulence. The polymorphic rhoptry protein kinase ROP18 was recently shown to determine the difference in virulence between type I and III strains by phosphorylating and inactivating the interferon-γ (IFNγ)-induced immunity-related GTPases (IRGs) that promote killing by disrupting the parasitophorous vacuole membrane (PVM) in murine cells. The polymorphic pseudokinase ROP5 determines strain differences in virulence through an unknown mechanism. Here we report that ROP18 can only inhibit accumulation of the IRGs on the PVM of strains that also express virulent ROP5 alleles. In contrast, specific ROP5 alleles can reduce IRG coating even in the absence of ROP18 expression and can directly interact with one or more IRGs. We further show that the allelic combination of ROP18 and ROP5 also determines IRG evasion and virulence of strains belonging to other lineages besides types I, II and III. However, neither ROP18 nor ROP5 markedly affect survival in IFNγ-activated human cells, which lack the multitude of IRGs present in murine cells. These findings suggest that ROP18 and ROP5 have specifically evolved to block the IRGs and are unlikely to have effects in species that do not have the IRG system, such as humans. PMID:22761577

  18. Mapping and identification of interferon gamma-regulated HeLa cell proteins separated by immobilized pH gradient two-dimensional gel electrophoresis

    DEFF Research Database (Denmark)

    Shaw, A.; Larsen, M.; Roepstorff, P.

    1999-01-01

    magnitude of IFN-gamma responsive genes has been reported previously. Our goal is to identify and map IFN-gamma-regulated HeLa cell proteins to the two-dimensional polyacrylamide gel electrophoresis with the immobilized pH gradient (IPG) two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) system...

  19. Serum levels of the interferon-gamma-inducing cytokine interleukin-18 are increased in individuals at high risk of developing type I diabetes

    DEFF Research Database (Denmark)

    Nicoletti, F; Conget, I; Di Marco, R

    2001-01-01

    and thought to be involved in its pathogenesis. Because increased production of IFN-gamma could be secondary to a dysregulated synthesis of IL-18, we compared the circulating levels of IL-18 in patients with newly diagnosed Type I diabetes with those of non-diabetic first-degree relatives and healthy control...

  20. Interferon alpha treatment stimulates interferon gamma expression in type I NKT cells and enhances their antiviral effect against hepatitis C virus.

    Science.gov (United States)

    Miyaki, Eisuke; Hiraga, Nobuhiko; Imamura, Michio; Uchida, Takuro; Kan, Hiromi; Tsuge, Masataka; Abe-Chayama, Hiromi; Hayes, C Nelson; Makokha, Grace Naswa; Serikawa, Masahiro; Aikata, Hiroshi; Ochi, Hidenori; Ishida, Yuji; Tateno, Chise; Ohdan, Hideki; Chayama, Kazuaki

    2017-01-01

    Interferon (IFN) inhibits hepatitis C virus (HCV) replication through up-regulation of intrahepatic IFN-stimulated gene expression but also through activation of host immune cells. In the present study, we analyzed the immune cell-mediated antiviral effects of IFN-α using HCV-infected mice. Urokinase-type plasminogen activator (uPA)-severe combined immunodeficiency (SCID) mice with transplanted human hepatocytes were infected with genotype 1b HCV and injected with human peripheral blood mononuclear cells (PBMCs). IFN-α treatment following human PBMC transplantation resulted in a significant reduction in serum HCV RNA titers and a higher human CD45-positive mononuclear cell chimerism compared to mice without human PBMC transplantation. In mice with human PBMCs treated with IFN-α, serum concentrations of IFN-γ increased, and natural killer T (NKT) cells, especially type I NKT cells, produced IFN-γ. Mice in which IFN-γ signaling was blocked using antibody or in which transplanted PBMCs were depleted for type I NKT cells showed similar levels of anti-HCV effect compared with mice treated only with IFN-α. These results show that IFN-α stimulates IFN-γ expression in type 1 NKT cells and enhances the inhibition of HCV replication. We propose that type 1 NKT cells might represent a new therapeutic target for chronic hepatitis C patients.