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Flow cytometry as a tool to assess the effects of gamma radiation on the viability, growth and metabolic activity of fungal spores

Mesquita, N.; Portugal, A.; Piñar, G.; Loureiro, J.; Coutinho, A. P.; Trovão, J.; Nunes, I.; Botelho, M. L.; Freitas, H.
Fonte: Elsevier Ltd. Publicador: Elsevier Ltd.
Tipo: Artigo de Revista Científica
ENG
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Flow cytometry is often used for viability and vitality assessment in bacteria and yeasts. However, its application to the study of fungal spore development is uncommon, probably due to the difficulties in successfully staining these cells. In the current study, we used flow cytometry for the first time to assess the effects of a disinfection treatment on the survival, growth and metabolic activity of fungal spores (Penicillium chrysogenum, Aspergillus nidulans and Aspergillus niger) submitted to gamma radiation (0e15 kGy). The Forward and Side-Scatter parameters of the cytometer were used to assess the differences in size and complexity of particles. Furthermore, two fluorescent dyes were used: Propidium Iodide to assess the membrane integrity and spore viability, in a culture-independent procedure; and Dihydroethidium to measure the changes in metabolic activity of irradiated spores in their first 10 h of growth in a liquid culture medium. Our results support that flow cytometry is a valuable tool in assessing different biological parameters and biocide effects, as it allowed accurate determination of the viability, growth and metabolic activity of gamma-irradiated spores. The fluorescence of Propidium Iodide was 5e7 more intense in unviable spores. The Dihydroethidium fluorescence increase was associated with faster growth. Control and low radiation doses allowed the germination and growth of spores...

Comparison of flow cytometry and indirect immunofluorescence assay in the diagnosis and cure criterion after therapy of American tegumentary leishmaniasis by anti-live Leishmania (Viannia) braziliensis immunoglobulin G

Oliveira, Andresa Pereira de; Castro, Maria Carolina Accioly Brelaz de; Almeida, Amanda Ferreira de; Souza, Marina de Assis; Oliveira, Beatriz Coutinho de; Reis, Luiza de Campos; Goto, Hiro; Brito, Maria Edileuza Felinto de; Celeste, Beatriz Julieta; Mart
Fonte: Amsterdam Publicador: Amsterdam
Tipo: Artigo de Revista Científica
ENG
Relevância na Pesquisa
66.48%
The aim of this study was to compare the techniques of indirect immunofluorescence assay (IFA) and flow cytometry to clinical and laboratorial evaluation of patients before and after clinical cure and to evaluate the applicability of flow cytometry in post-therapeutic monitoring of patients with American tegumentary leishmaniasis (ATL). Sera from 14 patients before treatment (BT), 13 patients 1 year after treatment (AT), 10 patients 2 and 5 years AT were evaluated. The results from flow cytometry were expressed as levels of IgG reactivity, based on the percentage of positive fluorescent parasites (PPFP). The 1:256 sample dilution allowed us to differentiate individuals BT and AT. Comparative analysis of IFA and flow cytometry by ROC (receiver operating characteristic curve) showed, respectively, AUC (area under curve) = 0.8 (95% CI = 0.64–0.89) and AUC = 0.90 (95% CI = 0.75–0.95), demonstrating that the flow cytometry had equivalent accuracy. Our data demonstrated that 20% was the best cut-off point identified by the ROC curve for the flow cytometry assay. This test showed a sensitivity of 86% and specificity of 77% while the IFA had a sensitivity of 78% and specificity of 85%. The after-treatment screening, through comparative analysis of the technique performance indexes...

Validação da metodologia de citometria de fluxo para avaliação da contagem bacteriana do leite cru; Evaluation of flow cytometry as a method for total bacterial count of raw milk

Cassoli, Laerte Dagher
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 16/08/2005 PT
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O objetivo do trabalho foi avaliar a utilização da metodologia de citometria de fluxo na determinação da contagem bacteriana total (CBT) do leite cru. No primeiro estudo foi avaliado o efeito da temperatura de armazenamento, da idade da amostra e do tipo conservante sobre a CBT. Também foi estudada a possibilidade de se utilizar uma única amostra de leite para a realização das análises previstas na Instrução Normativa 51 (IN-51). Foram testadas, três temperaturas de armazenamento (0oC – congelado, 7oC – resfriamento e 24 oC – ambiente), três conservantes (bronopol, azidiol e sem conservante) e quatro tempos entre a coleta e a análise (idade da amostra) (um (D1), três (D3), cinco (D5) e sete (D7) dias). Foi considerado tratamento controle para análise de CBT, amostras refrigeradas, com azidiol e com um dia de idade. Para as análises de composição e CCS, o tratamento controle foram amostras refrigeradas, com bronopol e com um dia de idade. Os resultados indicaram que será necessária a coleta de duas amostras, uma destinada à determinação de CCS e composição, contendo bronopol e, outra, para CBT, contendo azidiol. A amostra para CBT poderá ser analisada em até sete dias após a coleta, desde que mantida sob refrigeração à 7ºC. Deve-se evitar o aquecimento ou o congelamento da amostra para CBT...

Citometria de fluxo de leucócitos sangüíneos de Phrynops geoffroanus (Schweigger, 1812) provenientes de ambientes poluídos: metodologia de isolamento e estimulação; Flow cytometry of blood leucocytes of Phrynops geoffroanus (Schweigger, 1812) from polluted environments: isolation and stimulation methodology

Genoy-Puerto, Elmer Alexander
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 08/07/2008 PT
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66.3%
O quelônio Phrynops geoffroanus aparece freqüentemente associado a cursos d'água poluídos, sem que, no entanto, sejam conhecidos aspectos que descrevam sua atividade imune celular frente situações adversas (efeitos antrópicos). A citometria de fluxo permite mensurar características estruturais e imunológicas de células em suspensão submetidas a um fluxo contínuo, sendo capaz de analisar inúmeros tipos celulares. Neste trabalho foi desenvolvida metodologia para estudar e avaliar a função celular de leucócitos sangüíneos do Phrynops geoffroanus através da fagocitose e burst oxidativo espontâneo e induzido utilizando a citometria de fluxo. Para definir a metodologia foram utilizadas 86 amostras sangüíneas de Phrynops geoffroanus encontrados em ambientes antrópicos do rio Piracicaba, ribeirão Piracicamirim e Fundação Parque Zoológico de São Paulo como grupo de comparação. Estes resultados foram complementados pelo perfil hematológico. O processamento incluiu transporte e conservação das amostras em RPMI 1640 Gibco® para posterior utilização de Ficoll-PaqueTM PLUS como agente separador entre leucócitos e hemácias, utilizando-se centrifugações refrigeradas (18°C) com acelerações e desacelerações graduais. Amostras com porcentagens de viabilidade inferiores a 90 % não foram utilizadas na realização dos estímulos: Zymosan A (Saccharomyces cerevisiae) Bio Particles®...

Estudo das alterações morfo-funcionais de espermatozóides bovinos submetidos à sexagem por meio da técnica de citometria de fluxo; Study of morpho-functions alterations in bovine spermatozoa submitted on sorting through flow cytometry technology

Tanno, Priscilla Harumi
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 14/09/2009 PT
Relevância na Pesquisa
66.27%
O objetivo deste estudo foi avaliar as alterações morfo-funcionais do sêmen bovino submetido à sexagem através da técnica de citometria de fluxo e comparar com o sêmen convencional e a diferença entre as subespécies. O sêmen foi obtido de seis touros, sendo três taurinos (Holandês preto e branco) e três zebuínos (Gir leiteiro), com quatro ejaculados de cada animal (n = 24). Cinco tratamentos foram realizados: sêmen convencional com diluidor L (Lagoa); sêmen convencional com diluidor S (Sexing) e sêmen sexado (nos tempos de zero, três e seis horas após a ejaculação para avaliar se existem alterações de membranas ao longo do tempo). Foram avaliados pela citometria de fluxo quanto à integridade de membrana plasmática e reação acrossomal (PI/FITC-PSA); peroxidação lipídica (C11-BODIPY581/591) e capacitação espermática através de análises da fosforilação da tirosina e aumento da fluidez e desorganização da membrana plasmática (Merocianina 540 e Yo-Pro1). Os efeitos de tratamentos foram avaliados por análises de variância (ANOVA), empregando-se o programa estatístico Stat-View® (SAS Institute, Inc. 1998, Cary, NC, USA). Foram considerados significativos os valores com P<0,05 e com tendência a significância (P<0...

Estabelecimento de um ensaio funcional de ligação de proteínas de merozoítas de Plasmodium vivax a eritrócitos humanos por citometria de fluxo; Establishment of a functional binding assay of Plasmodium vivax merozoite proteins to human erythrocytes by flow cytometry

Françoso, Katia Sanches
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 25/10/2012 PT
Relevância na Pesquisa
66.42%
A invasão de eritrócitos por merozoítas de Plasmodium é um processo bastante complexo em que várias proteínas do parasita interagem com receptores presentes na célula hospedeira. É nossa hipótese que o Antígeno 1 de Membrana Apical (AMA-1) e a Proteína 1 da Superfície do Merozoíta (MSP1) estejam envolvidas na adesão ao eritrócito, no momento da invasão. Sendo assim, o objetivo deste estudo foi avaliar a capacidade de ligação de proteínas de merozoítas de P. vivax a eritrócitos humanos utilizando um novo ensaio de citometria de fluxo e o clássico ensaio de citoaderência. Para tanto, a ligação das proteínas recombinantes correspondentes à região C-terminal de 19 kDa da MSP1 (PvMSP119), ao ectodomínio (PvAMA-1E) e aos domínios I-II e II de AMA-1 (PvAMA-1DI-II e PvAMA-1DII) foi avaliada por citometria de fluxo utilizando anticorpo fluorescente contra cauda de histidina. A proteína recombinante baseada na região II da Proteína Ligante de Duffy (PvDBP-RII) foi utilizada como controle positivo e a tioredoxina humana como controle negativo. Os dados foram expressos pela média da intensidade de fluorescência (MIF). Os resultados dos ensaios de citometria de fluxo confirmaram a ligação de PvDBP-RII a eritrocitos Duffy A (MIF=1997±405) e Duffy B (MIF= 5760±303). No entanto...

Correlação da intensidade de fluorescência com o resultado gestacional no exame de prova cruzada por citometria de fluxo em mulheres submetidas a imunização com leucócitos paternos = : Correlation of fluorescence intensity with pregnancy outcome in crossmatch test by flow cytometry in women undergoing immunization with paternal leucocytes; Correlation of fluorescence intensity with pregnancy outcome in crossmatch test by flow cytometry in women undergoing immunization with paternal leucocytes

Michele Cintra Vicentini
Fonte: Biblioteca Digital da Unicamp Publicador: Biblioteca Digital da Unicamp
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 17/12/2013 PT
Relevância na Pesquisa
66.27%
Introdução: O sucesso da gestação envolve mecanismos aloimunes, sendo o feto alogênico considerado um aloenxerto bem sucedido. A imunização com leucócitos paternos têm se apresentado como uma eficiente opção terapêutica para os casos de abortamento recorrente, induzindo a produção de anticorpos anti-paternos pelo sistema imune materno durante a gestação. Atualmente, o método de Prova Cruzada por citometria de fluxo (FCXM) vem sendo utilizada para auxílio na avaliação de pacientes com Aborto Espontâneo de Repetição (AER). Objetivos: Avaliar o resultado do FCMX em mulheres com antecedente de AER que engravidaram após tratamento com ILP de acordo com o resultado gestacional. Material e Métodos: 85 pacientes foram selecionadas para o estudo tendo como critérios de inclusão: mulheres com dois ou mais AER, FXCM inicial negativo contra os maridos e mulheres de AER primário. As mulheres divididas em 2 dois grupos 1) Sucesso Gestacional (SG) 2) Perda Gestacional (PG). Após um FCXM inicial negativo (FCXM pré) as pacientes foram imunizadas com doses de ILP e após 30 dias da última ILP um novo FCXM (FCXM pós) foi realizado, a mediana da intensidade de fluorescência (MIF) foi calculada nos 2 momentos e comparada nos 2 grupos. Resultados: Calculando a razão da Intensidade de Fluorescência (IF FCXM pós / IF FCXM pré) para linfócitos T e B analisamos que não houve diferença significativa entre os dois grupos SG e PG...

Assessment of brewing yeast age based on selective bud scar staining and subsequent fluorescence measurement by flow cytometry

Lehnert, Radek; Kuřec, Michal; Brányik, Tomáš; Mota, André; Teixeira, J. A.
Fonte: Universidade do Minho Publicador: Universidade do Minho
Tipo: Conferência ou Objeto de Conferência
Publicado em 06/05/2007 ENG
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To study the yeast cell aging in continuous beer fermentation processes has a considerable practical significance. Hence, an expeditious method of yeast age estimation was developed based on selective bud scar staining and subsequent fluorescence intensity measurement by flow cytometry. The calibration curve resulting from cytometric determination of average bud scar fluorescence intensities of cell populations with different average bud scar numbers showed a good correlation. The combination of flow cytometry with bud scar staining resulted in a method of rapid cell age estimation particularly suitable for continuous main fermentation systems with elevated risk of immobilized cell aging.; Grant Agency of the Czech Republic - Project 104/06/1418.

Stage-specific activity of potential antimalarial compounds measured in vitro by flow cytometry in comparison to optical microscopy and hypoxanthine uptake

Contreras,Carmen E; Rivas,María A; Domínguez,José; Charris,Jaime; Palacios,Mario; Bianco,Nicolás E; Blanca,Isaac
Fonte: Instituto Oswaldo Cruz, Ministério da Saúde Publicador: Instituto Oswaldo Cruz, Ministério da Saúde
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/03/2004 EN
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The evaluation of new antimalarial agents using older methods of monitoring sensitivity to antimalarial drugs are laborious and poorly suited to discriminate stage-specific activity. We used flow cytometry to study the effect of established antimalarial compounds, cysteine protease inhibitors, and a quinolone against asexual stages of Plasmodium falciparum. Cultured P. falciparum parasites were treated for 48 h with different drug concentrations and the parasitemia was determined by flow cytometry methods after DNA staining with propidium iodide. P. falciparum erythrocytic life cycle stages were readily distinguished by flow cytometry. Activities of established and new antimalarial compounds measured by flow cytometry were equivalent to results obtained with microscopy and metabolite uptake assays. The antimalarial activity of all compounds was higher against P. falciparum trophozoite stages. Advantages of flow cytometry analysis over traditional assays included higher throughput for data collection, insight into the stage-specificity of antimalarial activity avoiding use of radioactive isotopes.

The sample processing time interval as an influential factor in flow cytometry analysis of lymphocyte subsets

Santos,Ana Paula dos; Bertho,Álvaro Luiz; Martins,Reinaldo de Menezes; Marcovistz,Rugimar
Fonte: Instituto Oswaldo Cruz, Ministério da Saúde Publicador: Instituto Oswaldo Cruz, Ministério da Saúde
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/02/2007 EN
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The objective of this paper is to propose a protocol to analyze blood samples in yellow fever 17DD vaccinated which developed serious adverse events. We investigated whether or not the time between sample collection and sample processing could interfere in lymphocyte subset percentage, for it is often impossible to analyze blood samples immediately after collection due to transport delay from collection places to the flow cytometry facility. CD4+CD38+ T, CD8+CD38+ T, CD3+ T, CD19+ B lymphocyte subsets were analyzed by flow cytometry in nine healthy volunteers immediately after blood collection and after intervals of 24 and 48 h. The whole blood lysis method and gradient sedimentation by Histopaque were applied to isolate peripheral blood mononuclear cells for flow cytometry analyses. With the lysis method, there was no significant change in lymphocyte subset percentage between the two time intervals (24 and 48 h). In contrast, when blood samples were processed by Histopaque gradient sedimentation, time intervals for sample processing influenced the percentage in T lymphocyte subsets but not in B cells. From the results obtained, we could conclude that the whole blood lysis method is more appropriate than gradient sedimentation by Histopaque for immunophenotyping of blood samples collected after serious adverse events...

Trypan blue exclusion assay by flow cytometry

Avelar-Freitas,B.A.; Almeida,V.G.; Pinto,M.C.X.; Mourão,F.A.G.; Massensini,A.R.; Martins-Filho,O.A.; Rocha-Vieira,E.; Brito-Melo,G.E.A.
Fonte: Associação Brasileira de Divulgação Científica Publicador: Associação Brasileira de Divulgação Científica
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/04/2014 EN
Relevância na Pesquisa
66.38%
Dye exclusion tests are used to determine the number of live and dead cells. These assays are based on the principle that intact plasma membranes in live cells exclude specific dyes, whereas dead cells do not. Although widely used, the trypan blue (TB) exclusion assay has limitations. The dye can be incorporated by live cells after a short exposure time, and personal reliability, related to the expertise of the analyst, can affect the results. We propose an alternative assay for evaluating cell viability that combines the TB exclusion test and the high sensitivity of the flow cytometry technique. Previous studies have demonstrated the ability of TB to emit fluorescence when complexed with proteins. According to our results, TB/bovine serum albumin and TB/cytoplasmic protein complexes emit fluorescence at 660 nm, which is detectable by flow cytometry using a 650-nm low-pass band filter. TB at 0.002% (w/v) was defined as the optimum concentration for distinguishing unstained living cells from fluorescent dead cells, and fluorescence emission was stable for 30 min after cell treatment. Although previous studies have shown that TB promotes green fluorescence quenching, TB at 0.002% did not interfere with green fluorescence in human live T-cells stained with anti-CD3/fluorescein isothiocyanate (FITC) monoclonal antibody. We observed a high correlation between the percentage of propidium iodide+CD3/FITC+ and TB+CD3/FITC+ cells...

Platelet antibody detection by flow cytometry: an effective method to evaluate and give transfusional support in platelet refractoriness

Bub,Carolina Bonet; Martinelli,Beatriz Moraes; Avelino,Thayna Mendonca; Goncalez,Ana Claudia; Barjas-Castro,Maria de Lourdes; Castro,Vagner
Fonte: Associação Brasileira de Hematologia e Hemoterapia e Terapia Celular Publicador: Associação Brasileira de Hematologia e Hemoterapia e Terapia Celular
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/01/2013 EN
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66.34%
BACKGROUND: Immune platelet refractoriness is mainly caused by human leukocyte antigen antibodies (80-90% of cases) and, to a lesser extent, by human platelet antigen antibodies. Refractoriness can be diagnosed by laboratory tests and patients should receive compatible platelet transfusions. A fast, effective and low cost antibody-screening method which detects platelet human leukocyte/platelet antigen antibodies is essential in the management of immune platelet refractoriness. OBJECTIVE: The aim of this study was to evaluate the efficiency of the flow cytometry platelet immunofluorescence test to screen for immune platelet refractoriness. METHODS: A group of prospective hematologic patients with clinically suspected platelet refractoriness treated in a referral center in Campinas, SP during July 2006 and July 2011 was enrolled in this study. Platelet antibodies were screened using the flow cytometry platelet immunofluorescence test. Anti-human leukocyte antigen antibodies were detected by commercially available methods. The sensitivity, specificity and predictive values of the immunofluorescence test were determined taking into account that the majority of antiplatelet antibodies presented human leukocyte antigen specificity. RESULTS: Seventy-six samples from 32 female and 38 male patients with a median age of 43.5 years (range: 5-84 years) were analyzed. The sensitivity of the test was 86.11% and specificity 75.00% with a positive predictive value of 75.61% and a negative predictive value of 85.71%. The accuracy of the method was 80.26%. CONCLUSION: This study shows that the flow cytometry platelet immunofluorescence test has a high correlation with the anti-human leukocyte antigen antibodies. Despite a few limitations...

GenePattern flow cytometry suite

Spidlen, Josef; Barsky, Aaron; Breuer, Karin; Carr, Peter; Nazaire, Marc-Danie; Hill, Barbara Allen; Qian, Yu; Liefeld, Ted; Reich, Michael; Mesirov, Jill; Wilkinson, Peter; Scheuermann, Richard H; Sekaly, Rafick-Pierre; Brinkman, Ryan R
Fonte: BioMed Central Publicador: BioMed Central
Tipo: Artigo de Revista Científica
EN_US
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Background: Traditional flow cytometry data analysis is largely based on interactive and time consuming analysis of series two dimensional representations of up to 20 dimensional data. Recent technological advances have increased the amount of data generated by the technology and outpaced the development of data analysis approaches. While there are advanced tools available, including many R/BioConductor packages, these are only accessible programmatically and therefore out of reach for most experimentalists. GenePattern is a powerful genomic analysis platform with over 200 tools for analysis of gene expression, proteomics, and other data. A web-based interface provides easy access to these tools and allows the creation of automated analysis pipelines enabling reproducible research. Results: In order to bring advanced flow cytometry data analysis tools to experimentalists without programmatic skills, we developed the GenePattern Flow Cytometry Suite. It contains 34 open source GenePattern flow cytometry modules covering methods from basic processing of flow cytometry standard (i.e., FCS) files to advanced algorithms for automated identification of cell populations, normalization and quality assessment. Internally, these modules leverage from functionality developed in R/BioConductor. Using the GenePattern web-based interface...

A probabilistic approach for the evaluation of minimal residual disease by multiparameter flow cytometry in leukemic B-cell chronic lymphoproliferative disorders; Cytometry Part A

Pedreira, C. E.; Costa, E. S.; Quijano, S.; Almeida, J.; Fernandez, C.; Florez, J.; Barrena, S.; Lecrevisse, Q.; Van Dongen, J. J. M.; Orfao, A.
Fonte: Pontifícia Universidade Javeriana Publicador: Pontifícia Universidade Javeriana
Formato: 1141-1150
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Vol. 73A, No. 12; Multiparameter flow cytometry has become an essential tool for monitoring response to therapy in hematological malignancies, including B-cell chronic lymphoproliferative disorders (B-CLPD). However, depending on the expertise of the operator minimal residual disease (MRD) can be misidentified, given that data analysis is based on the definition of expert-based bidimensional plots, where an operator selects the subpopulations of interest. Here, we propose and evaluate a probabilistic approach based on pattern classification tools and the Bayes theorem, for automated analysis of flow cytometry data from a group of 50 B-CLPD versus normal peripheral blood B-cells under MRD conditions, with the aim of reducing operator-associated subjectivity. The proposed approach provided a tool for MRD detection in B-CLPD by flow cytometry with a sensitivity of 8 3 1025 (median of 2 3 1027). Furthermore, in 86% of BCLPD cases tested, no events corresponding to normal B-cells were wrongly identified as belonging to the neoplastic B-cell population at a level of 1027. Thus, this approach based on the search for minimal numbers of neoplastic B-cells similar to those detected at diagnosis could potentially be applied with both a high sensitivity and specificity to investigate for the presence of MRD in virtually all B-CLPD. Further studies evaluating its efficiency in larger series of patients...

Evaluation of the Anti-Tumoural and Immune Modulatory Activity of Natural Products by Flow Cytometry; Clínica flow cytometry - Emerging applications

Fiorentino, Susana; Quijano, Sandra; Hernández, John Fredy; Urueña, Claudia; Santander, Sandra Paola; Cifuentes, Claudia
Fonte: In Tech Publicador: In Tech
Formato: 91-106
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Flow cytometry has shown to be a very useful tool in the assessment of plant-derived compounds, having potential anti-tumour activity. Our group has focused in the study of fractions and isolated compounds with biological activity over tumour cells and regulatory immune cells, such as dendritic cells. Also we have implemented a flow cytometry screening tests for estimating the biological activity of plant-derived fractions. The present chapter will discuss important and useful facts to be taken into account for effectively carrying out biological assays in this area. We will start with recommendations about the production of plant fractions and continue with the biological assessment of cell lines or monocyte-derived dendritic cells. Since these protocols are constantly evolving, herein we describe our current experience.; Rijeka, Croatia

Comparison of flow cytometry and immunofluorescence microscopy for the detection of Giardia duodenalis in bovine fecal samples

Uehlinger, F.; Barkema, H.; O'Handley, R.; Parenteau, M.; Parrington, L.; VanLeeuwen, J.; Dixon, B.
Fonte: Amer Assoc Veterinary Laboratory Diagnosticians Inc Publicador: Amer Assoc Veterinary Laboratory Diagnosticians Inc
Tipo: Artigo de Revista Científica
Publicado em //2008 EN
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66.34%
The performance of flow cytometry (FC) was compared with immunofluorescence microscopy (IM) for detection of Giardia duodenalis in bovine feces. Samples from 36 adult dairy cows and 208 dairy calves were collected. Flow cytometry test characteristics were calculated using continuous, ordinal, and dichotomized results. Spearman correlation coefficients comparing the results of the 2 tests were 0.47 and 0.68 for cows and calves, respectively. Using IM as indicative of presence or absence of G. duodenalis cysts in each sample, likelihood ratios of FC results with 0, 1, and ≥2 gated events indicated that samples with 1 gated event were likely to be positive in the cows but not in the calves. Immunofluorescence microscopy detected G. duodenalis in 69.7% and 48.1% of cows and calves, respectively. When dichotomizing the FC results at a cutoff point of 1 or 2 gated events, 46.3% and 19.9% of the cow and 51.9% and 35.1% of the calf samples, respectively, were classified as G. duodenalis-positive. Relative to IM, the sensitivity in the cows was 0.59 and 0.28, respectively, and 0.76 and 0.64, respectively, in the calves. At a cut-off point of 1, 65.7% and 73.1% of the cow and calf samples, respectively, were correctly classified in FC, and at a cut-off point of 2...

Detection of PNH cells by flow cytometry, using multiparameter analysis

Rocha,Juliana Maria Camargos; Silva,Maria Luíza; Souza,Marcelo Eduardo de Lima; Murao,Mitiko; Araújo,Sérgio Schusterschitz da Silva; Santos,Silvana Maria Elói
Fonte: Sociedade Brasileira de Patologia Clínica; Sociedade Brasileira de Patologia; Sociedade Brasileira de Citopatologia Publicador: Sociedade Brasileira de Patologia Clínica; Sociedade Brasileira de Patologia; Sociedade Brasileira de Citopatologia
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/04/2014 EN
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66.34%
Introduction: The laboratory diagnosis of paroxysmal nocturnal hemoglobinuria (PNH), disease that is categorized by reduced synthesis of glycosylphosphatidylinositol (GPI) anchor, is based on the detection of blood cells deficient in GPI-anchored proteins by flow cytometry. PNH clones have been detected in patients with aplastic anaemia (AA) and myelodysplastic syndrome (MDS), with therapeutic implications. Objectives: To validate a sensitive assay for detection of GPI-anchored protein-deficient cells, by flow cytometry, and to analyze the clone frequency in AA and MDS patients. Methods: Samples from 20 AA patients, 30 MDS patients and 20 adult volunteers (control group) were analyzed using monoclonal antibodies to CD16, CD24, CD55 and CD59 (neutrophils); CD14 and CD55 (monocytes); CD55 and CD59 (erythrocytes); besides fluorescent aerolysin reagent (FLAER) (neutrophils and monocytes) and lineage markers. The proportions of PNH cells detected in neutrophils and monocytes, using different reagent combinations, were compared by analysis of variance (ANOVA) and Pearson's correlation. Results: PNH cells were detected in five (25%) AA patients, and the proportions of PNH cells varied from 0.14% to 94.84% of the analyzed events. PNH cells were not detected in the MDS patients. However...

Clinical role of flow cytometry in redefining bone marrow involvement in diffuse large B-cell lymphoma (DLBCL) - a new perspective

Talaulikar, Dipti; Shadbolt, Bruce; Bell, J; Khan, K; Dahlstrom, Jane; McDonald, A; Pidcock, Michael; Broomfield, Amy
Fonte: Wiley-Blackwell Publicador: Wiley-Blackwell
Tipo: Artigo de Revista Científica
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66.35%
Aims: The clinical role of flow cytometry in staging bone marrow in diffuse large B-cell lymphoma (DLBCL), especially its impact on outcome, remains uncertain. The aim was to determine the contribution of flow cytometry to conventional staging, and to stu

Using flow cytometry for counting natural planktonic bacteria and understanding the structure of planktonic bacterial communities

Gasol, Josep M.; Del Giorgio, Paul A.
Fonte: CSIC - Instituto de Ciencias del Mar (ICM) Publicador: CSIC - Instituto de Ciencias del Mar (ICM)
Tipo: Artículo Formato: 695030 bytes; application/pdf
ENG
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Publicación online disponible en: http://www.icm.csic.es/scimar/index.php; Flow cytometry is rapidly becoming a routine methodology in aquatic microbial ecology. The combination of simple to use bench-top flow cytometers and highly fluorescent nucleic acid stains allows fast and easy determination of microbe abundance in the plankton of lakes and oceans. The different dyes and protocols used to stain and count planktonic bacteria as well as the equipment in use are reviewed, with special attention to some of the problems encountered in daily routine practice such as fixation, staining and absolute counting. One of the main advantages of flow cytometry over epifluorescence microscopy is the ability to obtain cell-specific measurements in large numbers of cells with limited effort. We discuss how this characteristic has been used for differentiating photosynthetic from non-photosynthetic prokaryotes, for measuring bacterial cell size and nucleic acid content, and for estimating the relative activity and physiological state of each cell. We also describe how some of the flow cytometrically obtained data can be used to characterize the role of microbes on carbon cycling in the aquatic environment and we prospect the likely avenues of progress in the study of planktonic prokaryotes through the use of flow cytometry.; This work has been supported by grants MAS3-CT95- 0016 (MEDEA) and MAS3-CT97-0154 (MIDAS).; Peer reviewed

Optimization of flow cytometry assays for the detection of injured foodborne pathogenic bacteria

Subires Orenes, Alicia
Fonte: [Barcelona] : Universitat Autònoma de Barcelona, Publicador: [Barcelona] : Universitat Autònoma de Barcelona,
Tipo: Tesis i dissertacions electròniques; info:eu-repo/semantics/doctoralThesis; info:eu-repo/semantics/publishedVersion
Publicado em //2015 ENG
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Algunos tratamientos de elaboración y conservación de los alimentos lesionan las bacterias, de manera que pierden la capacidad para multiplicarse y no son detectadas por métodos de cultivo convencionales. La citometría de flujo es una técnica de análisis independiente del cultivo de los microorganismos, usada ampliamente para evaluar de manera rápida el estado fisiológico de células bacterianas individuales, siendo la integridad de membrana unos de los indicadores más habituales debido a su importancia en la supervivencia de éstas. Sin embargo, los resultados obtenidos mediante citometría de flujo se ven afectados por la presencia de partículas de las muestras alimentarias. En esta tesis, se evaluaron estrategias para desarrollar un ensayo por citometría de flujo basado en el uso del yoduro de propidio (YP), fluorocromo que no atraviesa las membranas intactas, combinado con un fluorocromo que penetra las membranas intactas, para detectar y cuantificar con precisión bacterias patógenas lesionadas. En el primer experimento, se ajustaron las concentraciones y las ratios de combinaciones de YP con SYTO 9, SYTO 24 y SYTO BC para mejorar la resolución de poblaciones de células sanas y muertas de Escherichia coli O157:H7...