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Antígeno excreción-secreción de Fasciola hepatica: ultrafiltración y aplicación en inmunodiagnóstico 38
Colmenares,Cecilia; Méndez,Liber; Díaz-Bello,Zoraida; Alarcón de Noya,Belkisyolé.
La baja sensibilidad de la coprología en el diagnóstico de la fasciolosis humana ha motivado el desarrollo del inmunodiagnóstico. El antígeno de excreción-secreción de adultos de Fasciola hepatica (AFhES) en ELISA es adecuado para el screening, aunque sobrestima la prevalencia; cuando se utiliza en Western blot (WB) no muestra un buen reconocimiento de sus componentes. Para lograr una mejor preparación, se ultrafiltró el antígeno a través de membranas YM de 10, 30 y 50 kDa. Los retenidos (R) se usaron en ELISA y WB. La mayor discriminación entre positivos y negativos en ELISA y la mejor resolución en el reconocimiento al antígeno en WB, se logró con la fracción R50. Se destacan las moléculas de 9, 14, 65 kDa y la región alrededor de 27 kDa, detectadas con...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Fasciolosis; Antígeno; Ultrafiltración; Inmunodiagnóstico; ELISA; Western blot.
Ano: 2007 URL: http://www.scielo.org.ar/scielo.php?script=sci_arttext&pid=S0325-29572007000200010
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Characterization of Mycoplasma penetrans and Mycoplasma fermentans immunodominant proteins 58
Bruder,Juliana; Godry,Renata Cristina; Takei,Kioko; Cunha,Regina Ayr Florio da.
Mycoplasmas are a heterogeneous group of the smallest organisms capable of self replication and are known to cause many detrimental diseases in both animals and humans. These wall-less prokaryotes are enveloped by a lipoprotein membrane and their small genomes are sufficient to synthesize molecules required for growth and self-replication. Among sixteen species isolated from humans, Mycoplasma pneumoniae, an agent of primary atypical pneumonia, and the urogenital tract species Mycoplasma hominis,Ureaplasma urealyticum and Ureaplasma parvum have been confirmed to be pathogenic. Mycoplasma penetrans and Mycoplasma fermentans, which are species associated with HIV, have been investigated mainly in research laboratories. In this study we have characterized...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Mycoplasma penetrans; Mycoplasma fermentans; LAMP-proteins; ELISA; Western blot.
Ano: 2005 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822005000200007
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Comparación del algoritmo nuevo con el algoritmo convencional para diagnóstico de VIH en Bolivia 38
Martínez-Oliva,Brenda Gisela; Montaño-Valenzuela,Karen Jimena; Rodriguez-Herbas,Patricia; Flores-León,Amilcar Alejandr; Grados-Torrez,Ricardo Enrique.
En Bolivia, el algoritmo diagnóstico convencional para VIH utiliza como prueba confirmatoria el Western blot; debido a dificultades con el costo, tiempo y centralización del diagnóstico confirmatorio, el Programa Nacional ITS/VIH/SIDA-HV propuso un algoritmo nuevo que utiliza solo dos pruebas rápidas y ELISA Ag/Ac. Sesenta muestras de suero fueron procesadas con ambos algoritmos y se determinó el resultado verdadero de las muestras de acuerdo con ambos algoritmos y la carga viral. La sensibilidad, especificidad, valores predictivos, índice Kappa y chi-cuadrado de Pearson fueron calculados. El algoritmo convencional reportó: sensibilidad 96,3%, especificidad 100%, valor predictivo positivo 100% y valor predictivo negativo 97% el algoritmo nuevo presentó...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Diagnóstico de VIH; Prueba rápida; Western blot; Carga viral.
Ano: 2018 URL: http://www.scielo.org.ar/scielo.php?script=sci_arttext&pid=S0325-29572018000200007
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Crude alkaloid extract of Rhazya stricta inhibits cell growth and sensitizes human lung cancer cells to cisplatin through induction of apoptosis 74
Elkady,Ayman I..
There is an urgent need to improve the clinical management of non-small cell lung cancer (NSCLC), one of the most frequent causes of cancer-related deaths in men and women worldwide. Rhazya stricta, an important medicinal plant used in traditional Oriental medicine, possesses anti-oxidant, anti-carcinogenic and free radical scavenging properties. This study was done to explore the potential anticancer activity of a crude alkaloid extract of R. stricta (CAERS) against the NSCLC line A549. CAERS markedly suppressed the growth of A549 cells and considerably enhanced the anti-proliferative potential of cisplatin. CAERS-mediated inhibition of A549 cell growth correlated with the induction of apoptosis that was accompanied by numerous morphological changes, DNA...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Apoptosis; Lung cancer; Medicinal plant; RT-PCR; Western blot.
Ano: 2013 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1415-47572013000100003
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Detection and coat protein gene characterization of an isolate of Grapevine virus B from corky bark-affected grapevines in Southern Brazil 102
NICKEL,OSMAR; FAJARDO,THOR V. M.; ARAGÃO,FRANCISCO J. L.; CHAGAS,CÉSAR M.; KUHN,GILMAR B..
An isolate of Grapevine virus B (GVB), obtained by indexing Vitis labrusca and V. vinifera grapevines on the indicator LN33, was transmitted mechanically to several Nicotiana species. The virus was partially purified from N. cavicola and the coat protein estimated at 23 kDa by SDS-PAGE. In negatively stained leaf extracts of experimentally inoculated N. cavicola and N. occidentalis, flexuous particles with cross banding were observed, predominantly measuring 750-770 x 12 nm, with a modal length of 760 nm. Decoration indicated a clear, positive reaction against AS-GVB. In DAS-ELISA, GVB was detected in N. cavicola and grapevine extracts, and Western blots showed homologous and cross reaction of GVB and GVA antisera with GVB coat protein. Using specific...
Tipo: Info:eu-repo/semantics/article Palavras-chave: GVB; Vitis spp.; Vitivirus; Electron microscopy; ELISA; Western blot; GVA.
Ano: 2002 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-41582002000300007
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Effects of single and double infections with Potato virus X and Tobacco mosaic virus on disease development, plant growth, and virus accumulation in tomato 102
BALOGUN,OLUSEGUN S.; XU,LEIXIN; TERAOKA,TOHRU; HOSOKAWA,DAIJIRO.
The tomato cv. Fukuju nº. 2 was used for studying the effect of single and double infections with Potato virus X (PVX) and Tobacco mosaic virus (TMV). Mixed infection resulted in a synergistic increase of disease severity, where more growth reduction was seen with simultaneous inoculations than with sequential inoculations at four-day intervals. At five and 12 days post-inoculation, the increased severity of the disease coincided with enhancement of virus accumulation in the rapidly expanding upper leaves. The PVX concentration in leaves nº 5 to 7 of doubly infected plants was three to six fold that of singly infected ones, as determined by DAS-ELISA. Mixed infection with the L strain led to higher enhancement of PVX than with the TMV-L11A strain. The...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Synergism; Western blot; Northern hybridization; DAS-ELISA; Virus enhancement.
Ano: 2002 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-41582002000300001
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Evidence of an endogenous lectin receptor in seeds of the legume Cratylia floribunda 61
Ramos,Márcio Viana; Bomfim,Liezelotte Rezende; Bandeira,; Debray,Henri.
Cratylia floribunda seeds were ground and the clean crude saline extract was fractionated into albumin, globulin, prolamin, acidic and basic glutelin protein fractions. These protein fractions were examined for the presence of an endogenous lectin receptor by SDS-polyacrylamide gel electrophoresis, western blot, affinity chromatography on a Sepharose 4B-Cratylia floribunda (CFL) lectin column and kinetic analysis in real time by surface plasmon resonance (SPR). Prolamin was the richest protein fraction although very poor in haemagglutinating activity. Basic glutelin was far the less interesting fraction for lectin activity and protein content, even though this fraction contains considerable amounts of carbohydrates. Lectin was present in all protein...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Protein-carbohydrate interaction; Surface plasmon resonance; Western blot.
Ano: 2002 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1677-04202002000300003
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Expressão em Escherichia coli da proteína capsidial do Watermelon mosaic virus e produção de anti-soro 102
Barbieri,Manuela R.; Carvalho,Murilo G. de; Zambolim,Eunize M.; Zerbini,F. Murilo.
Um anti-soro policlonal específico para Watermelon mosaic virus (WMV) foi produzido por meio da imunização de coelhos com proteína capsidial purificada, expressa in vitro em células de Escherichia coli. O gene cp foi amplificado via RT-PCR utilizando oligonucleotídeos específicos, a partir de RNA viral extraído de preparações virais concentradas. O fragmento amplificado foi clonado em pBLUESCRIPT KS+ e completamente seqüenciado para confirmação de sua identidade e integridade. Em seguida, o fragmento foi subclonado no vetor de expressão pRSET-A. Plasmídeos recombinantes foram utilizados para a expressão da proteína capsidial em E. coli BL21::DE3. A proteína foi purificada por meio de cromatografia de afinidade em coluna de Ni+-NTA, a partir de proteínas...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Cucurbitáceas; ELISA; Imunodifusão; Western blot.
Ano: 2004 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-41582004000200017
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Expression of a Haemonchus contortus cysteine protease in the baculovirus system 69
Miranda-Miranda,Estefan; Murillo-Sánchez,María Hortensia; Cossío-Bayúgar,Raquel.
A Haemonchus contortus recombinant Cysteine Protease (CP) was expressed in the baculovirus system. The CP gene was isolated by PCR from H. contortus cDNA, the PCR amplicon was cloned downstream to the polihedrin promoter within a bacterial expression vector, Sf9 insect cells were used for simultaneous co-transfection with the CP-vector and baculovirus naked DNA, which originated recombinant viruses by homologous recombination capable to express recombinant CP in an insect cell culture. A recombinant protease was identified as a fusion protein with a Ni lithium affinity 6XHis group. Recombinant CP was purified by affinity chromatography to obtain active recombinant protease identified by H. contortus experimentally infested ovine sera on a western blot as a...
Tipo: Journal article Palavras-chave: Haemonchosis; Molecular cloning; Recombinant protease; Synthetic substrates; Western blot.
Ano: 2008 URL: http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582008000200007
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Feline immunodeficiency virusinfection: a comparative study of different diagnostic techniques 45
Mortola,E.; Oliva,G.; Risso,M.; Pecoraro,M.; Venturini,M.C..
This study evaluated an indirect immunofluorescence assay (IFA) to detect feline immunodeficiency virus infection (FIV) antibody in a comprehensive epidemiological survey of FIV in Argentina. IFA modified in our laboratory, was compared with two other immunoassays, western blot (WB) and a sandwich immunochromatographic commercial kit (SI), and also with a direct polymerase chain reaction (PCR) method that detects proviral DNA. IFA showed to be a test with high sensitivity and specificity, and could be useful as a diagnostic tool in epidemiological studies. The presence of a low percentage of results with non-specific reactivity in the IFA could be resolved with further testing or use of an alternative method.
Tipo: Info:eu-repo/semantics/article Palavras-chave: Feline immunodeficiency virus; Immunofluorescence assay; Western blot; Polymerase chain reaction.
Ano: 2004 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0102-09352004000100003
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Heterologous expression of staphylococcal enterotoxin B (seb) gene for antibody production 69
Vrat Kamboj,Dev; Nema,Vijay; Kumar Pandey,Arun; Kumar Goel,Ajay; Singh,Lokendra.
Staphylococcal food poisoning (SFP) is caused by the members of superantigen family called staphylococcal enterotoxins (SEs). About 20 different types of SEs are produced by Staphylococcus aureus out of which type A (SEA), B (SEB), C (SEC) and D (SED) are commonly implicated in SFP. Among these, SEB is the most potent toxin and has also gained the status of biological warfare (BW) agent. Therefore, detection of SEB is of utmost importance. Any immunological detection system for SEB requires specific and sensitive antibodies which inturn depends on the purity of the SEB. In the present investigation, seb gene of S. aureus was cloned and expressed in E. coli along with biotin as fusion partner to facilitate the purification process. The yield of purified...
Tipo: Journal article Palavras-chave: Affinity purification; Biotin fusion; ELISA; Gene cloning; SEB; Western blot.
Ano: 2006 URL: http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582006000500010
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Humoral response to low molecular weight antigens of Mycobacterium tuberculosis by tuberculosis patients and contacts 56
Beck,S.T.; Leite,O.M.; Arruda,R.S.; Ferreira,A.W..
Much effort has been devoted to the identification of immunologically important antigens of Mycobacterium tuberculosis and to the combination of target antigens to which antibodies from serum of tuberculous patients could react specifically. We searched for IgG antibodies specific for antigens of 45 to 6 kDa obtained after sonication of the well-characterized wild M. tuberculosis strain in order to detect differences in the antibody response to low molecular weight antigens from M. tuberculosis between patients with pulmonary tuberculosis and contacts. Specific IgG antibodies for these antigens were detected by Western blot analysis of 153 serum samples collected from 51 patients with confirmed pulmonary tuberculosis. Three samples were collected from each...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Western blot; Profile; Tuberculosis; Antigens.
Ano: 2005 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2005000400013
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Identification of the autophagy pathway in a mollusk bivalve, Crassostrea gigas 5
Picot, Sandy; Faury, Nicole; Arzul, Isabelle; Chollet, Bruno; Renault, Tristan; Morga, Benjamin.
The Pacific oyster, Crassostrea gigas, is a mollusk bivalve commercially important as a food source. Pacific oysters are subjected to stress and diseases during culture. The autophagy pathway is involved in numerous cellular processes, including responses to starvation, cell death, and microorganism elimination. Autophagy also exists in C. gigas, and plays a role in the immune response against infections. Although this process is well-documented and conserved in most animals, it is still poorly understood in mollusks. To date, no study has provided a complete overview of the molecular mechanism of autophagy in mollusk bivalves. In this study, human and yeast ATG protein sequences and public databases (Uniprot and NCBI) were used to identify protein members...
Tipo: Text Palavras-chave: Autophagy; Autophagy related; Crassostrea gigas; Data mining; Immunohistochemistry; Real-time PCR; Western blot.
Ano: 2020 URL: https://archimer.ifremer.fr/doc/00606/71785/70270.pdf
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Immunohistochemical and molecular expression of laminin-332 gamma-2 chain in canine mammary tumors 45
Zuccari,D.A.P.C; Castro,R; Jardim,B.V; Mancini,U.M; Polachini,G.M.
Forty-eight cases of canine mammary cancer were investigated to evaluate the immunohistochemical distribution of the γ2 chain of laminin-332. Tumor cells were compared to a pool of normal mammary tissues using quantitative RT-PCR. The western blot was performed in eight tumor samples as complementary test to evaluate protein integrity. Immunohistochemistry experiments showed negative, focal, and weak expression of laminin-332 γ2 in tumors with the worst prognosis. Quantitative PCR revealed downregulation of the gene in 27 (56.2%) of the animals. Out of the 16 dogs with γ2 chain overexpression, seven were still alive. The western blot results showed bands generation of 36, 50, and 98kDa, suggesting degradation of laminin-332 γ2 in malignant tumors. The...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Dog; Adhesion molecule; Cancer; Western blot; RT-PCR.
Ano: 2011 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0102-09352011000100005
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Indirect ELISA for diagnosis of Brucella ovis infection in rams 45
França,S.A.; Mol,J.P.S.; Costa,E.A.; Silva,A.P.C.; Xavier,M.N.; Tsolis,R.M.; Reis,J.K.P.; Paixão,T.A.; Santos,R.L..
Brucella ovis is a major cause of epididymitis in sexually mature rams, resulting in subfertility, infertility, and economic losses for the sheep industry worldwide. The aim of this study was to develop an indirect ELISA (iELISA) using recombinant proteins, namely rBoP59 and rBP26, as antigens for serological diagnosis of B. ovisinfection. The BoP59 and BP26 recombinant proteins were expressed in E. coli and purified by affinity chromatography. Antigenicity was tested by Western blot and iELISA. Standardization of iELISA was performed with 500ng and 1µg BoP59 and rBP26 per well, testing serum from uninfected and experimentally infected rams. rBP26 was effective in distinguishing positive from negative rams. The rBP26 iELISA developed in this study is the...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Brucella ovis; Brucellosis; ELISAi; BP26; Western blot.
Ano: 2014 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0102-09352014000601695
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PPM1B and P-IKKβ expression levels correlated inversely with rat gastrocnemius atrophy after denervation 56
Wei,Jian; Liang,Bing-Sheng.
Activated inhibitor of nuclear factor-κB kinase β (IKKβ) is necessary and sufficient for denervated skeletal muscle atrophy. Although several studies have shown that Mg2+/Mn2+-dependent protein phosphatase 1B (PPM1B) inactivated IKKβ, few studies have investigated the role of PPM1B in denervated skeletal muscle. In this study, we aim to explore the expression and significance of PPM1B and phosphorylated IKKβ (P-IKKβ) during atrophy of the denervated gastrocnemius. Thirty young adult female Wistar rats were subjected to right sciatic nerve transection and were sacrificed at 0 (control), 2, 7, 14, and 28 days after denervation surgery. The gastrocnemius was removed from both the denervated and the contralateral limb. The muscle wet weight ratio was...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Muscular atrophy; Denervation; PPM1B; P-IKKβ; IKKβ; RT-PCR; Western blot.
Ano: 2012 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2012000800004
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Produção de antígeno e separação da proteína p28 por microfiltragem seriada para sorodiagnóstico da artrite encefalite caprina por ensaio imunoenzimático 45
Alves,L.A.O.; Teixeira,M.F.S.; Pinheiro,A.A.; Pinheiro,R.R.; Dias,R.P.; Brito,R.L.L.; Lopes Júnior,C.A.F.; Bezerra Júnior,R.Q.; Azevedo,D.A.A..
Este estudo teve como objetivo produzir um antígeno (Ag) a partir de cultura de células de membrana sinovial caprina (MSC) infectadas com o vírus de artrite encefalite caprina (CAEV), pela técnica de microfiltração seriada, substituindo a ultracentrifugação em colchão de sacarose (UCCS) para utilização em ELISA indireto (ELISA-i). Amostras de 188 soros caprinos, que previamente foram testados pelo Western blot (WB) com Ag UCCS, foram submetidas à análise pelo ELISA-i com o novo antígeno produzido, que mostrou concordância de 92% em relação ao antígeno UCCS. A sensibilidade e a especificidade do ELISA em relação ao WB foram de 95,6% e 88,5%, respectivamente. A nova técnica, criada a partir de microfiltrações, mostrou-se efetiva e de baixo custo para o...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Caprino; CAEV; ELISA; Lentivírus de pequenos ruminantes; Western blot.
Ano: 2012 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0102-09352012000400021
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Production of polyclonal antisera to manganese superoxide dismutase expressed in downy mildew resistant pearl millet and its application for immunodiagnosis 69
Urs Babitha,Mysore Pandurangaraj; Madhu,Devaiah; Sripathi P,Harishchandra; Shekar Shetty,Hunthrike.
Level of superoxide dismutase (SOD) in the seedlings of downy mildew resistant pearl millet genotype (IP 18293) increased 3-fold upon inoculation with Sclerospora graminicola. Mn-SOD was purified from pearl millet by ion-exchange and gel filtration chromatography. Polyclonal antibody was raised in rabbit against Mn-SOD of pearl millet. Anti-Mn SOD had an antibody titer of 1:20,000. ELISA results revealed a 1.5-fold increase in pure Mn-SOD preparation compared to the crude preparation. The antibody reactivity as detected by ELISA revealed that Mn-SOD is higher at 4 hrs post-inoculation in resistant pearl millet seedlings. The reactivity also showed the Mn-SOD is more in roots of inoculated resistant seedlings. SOD reactivity was also determined by DIBA...
Tipo: Journal article Palavras-chave: DIBA; Downy mildew; ELISA; Immunofluorescence; Manganese superoxide dismutase; Pearl millet; Sclerospora graminicola; Western blot.
Ano: 2006 URL: http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582006000200007
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Production of polyclonal antisera using recombinant coat proteins of Grapevine leafroll-associated virus 2 and Grapevine virus B 86
Radaelli,Paula; Fajardo,Thor Vinícius Martins; Nickel,Osmar; Eiras,Marcelo; Pio-Ribeiro,Gilvan.
The objective of this work was to produce and characterize specific antisera against Brazilian isolates of Grapevine leafroll-associated virus 2 (GLRaV-2) and Grapevine virus B (GVB), developed from expressed coat proteins (CPs) in Escherichia coli, and to test their possible use for the detection of these two viruses in diseased grapevines. The coat protein (CP) genes were RT-PCR-amplified, cloned and sequenced. The CP genes were subsequently subcloned, and the recombinant plasmids were used to transform E. coli cells and express the coat proteins. The recombinant coat proteins were purified, and their identities were confirmed by SDS-PAGE and Western blot and used for rabbit immunizations. Antisera raised against these proteins were able to recognize the...
Tipo: Info:eu-repo/semantics/article Palavras-chave: Vitis; GLRaV-2; GVB; Indirect ELISA; Recombinant protein; Western blot.
Ano: 2008 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-204X2008001000020
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Radiation dose determines the method for quantification of DNA double strand breaks 42
BULAT,TANJA; KETA,OTILIJA; KORIĆANAC,LELA; ŽAKULA,JELENA; PETROVIĆ,IVAN; RISTIĆ-FIRA,ALEKSANDRA; TODOROVIĆ,DANIJELA.
ABSTRACT Ionizing radiation induces DNA double strand breaks (DSBs) that trigger phosphorylation of the histone protein H2AX (γH2AX). Immunofluorescent staining visualizes formation of γH2AX foci, allowing their quantification. This method, as opposed to Western blot assay and Flow cytometry, provides more accurate analysis, by showing exact position and intensity of fluorescent signal in each single cell. In practice there are problems in quantification of γH2AX. This paper is based on two issues: the determination of which technique should be applied concerning the radiation dose, and how to analyze fluorescent microscopy images obtained by different microscopes. HTB140 melanoma cells were exposed to γ-rays, in the dose range from 1 to 16 Gy. Radiation...
Tipo: Info:eu-repo/semantics/article Palavras-chave: ApoTome software; AxioImagerA1 microscope; Immunofluorescence microscopy; Western blot; ΓH2AX.
Ano: 2016 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652016000100127
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