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1.
Mem. Inst. Oswaldo Cruz ; 115: e200214, 2020. tab, graf
Article in English | LILACS, SES-SP | ID: biblio-1135280

ABSTRACT

BACKGROUND Chagas disease, resulting from Trypanosoma cruzi infections, continues to be a health concern mainly in Latin American countries where the parasite is endemic. The laboratory diagnosis of a chronic infection is determined through serological assays for antibodies against T. cruzi and several tests are available that differ in key components, formats and methodologies. To date, no single test meets the criteria of a gold standard. The situation is further complicated by the difficulties associated with performance comparisons between different immunoassays or methodologies executed at different times and geographical areas. OBJECTIVE To improve the diagnosis of Chagas disease, the WHO coordinated the development of two International Biological Reference Standards for antibodies against anti-T. cruzi: NIBSC 09/186 and NIBSC 09/188 that respectively represent geographical regions with the highest prevalence of TcII and TcI lineages of the parasite. METHODS The principle goal of this study was to verify the behavior of these standards when assayed by several commercially available serological tests that employ different methods to capture and detect human anti-T. cruzi antibodies. FINDINGS AND MAIN CONCLUSIONS The results reinforce the recommendation that these standards be considered for performance evaluations of commercialised immunoassays and should be an integral step in the development of new test components or assay paradigms.


Subject(s)
Humans , Trypanosoma cruzi/isolation & purification , Serologic Tests/standards , Chagas Disease/diagnosis , Reference Standards , Trypanosoma cruzi/immunology , World Health Organization , Immunoassay/methods , Serologic Tests/methods , Antibodies, Protozoan/blood , Chagas Disease/parasitology
2.
Braz. j. microbiol ; 49(4): 885-890, Oct.-Dec. 2018. tab, graf
Article in English | LILACS | ID: biblio-974312

ABSTRACT

ABSTRACT In this study, the performance of the "RESIST-3 O.K.N. K-SeT" (Coris BioConcept, Gembloux, Belgium) immunochromatographic assay was evaluated in 132 Klebsiella pneumoniae comprising 102 carbapenem resistant and 30 carbapenem susceptible isolates. Genotypically known isolates of Gram negative bacteria (n = 22) including various species were also tested by the assay as controls. The isolates tested by the immunochromatographic assay and also were run PCR for bla KPC, bla IMP, bla VIM, bla NDM, and bla OXA-48. The rates of bla NDM, bla OXA-48, and bla KPC in carbapenem resistant isolates were found at 52.9%, 39.2%, and 2.0%, respectively. Both bla NDM and bla OXA-48 were found in six (5.9%) isolates. The results of the assay showed 100% concordance with those obtained by PCR in 132 K. pneumoniae. The agreement between the two methods was found to be identical at the isolate level. The assay also correctly detected all genotypically known isolates of Escherichia coli, Serratia marcescens, Citrobacter freundii, Enterobacter cloacae, K. pneumoniae carrying bla KPC, bla NDM, and/or bla OXA-48. On the other hand, the assay did not exhibit any cross-reaction in control isolates harboring bla IMP and bla VIM. We conclude that the RESIST-3 O.K.N. K-SeT is a reliable, rapid, and user friendly test and we recommend it for routine diagnostic laboratories.


Subject(s)
Humans , Bacterial Proteins/analysis , beta-Lactamases/analysis , Klebsiella Infections/microbiology , Immunoassay/methods , Klebsiella pneumoniae/enzymology , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Turkey , beta-Lactamases/metabolism , Carbapenems/pharmacology , Polymerase Chain Reaction , Klebsiella pneumoniae/isolation & purification , Klebsiella pneumoniae/drug effects , Klebsiella pneumoniae/chemistry , Anti-Bacterial Agents/pharmacology
3.
Rev. chil. endocrinol. diabetes ; 11(3): 103-107, jul. 2018. tab, ilus
Article in Spanish | LILACS | ID: biblio-915200

ABSTRACT

Abstract: Thyroid function is assessed by measuring thyrotropin and free and total thyroid hormone concentrations. There are interferences with the results of immunoassays that can lead to an incorrect diagnosis, of which the most frequent are the binding of thyroid hormones to heterophile antibodies, rheumatoid factor, anti-Ruthenium antibodies, the intake of biotin and anti-streptavidin antibodies. We present three cases of clinically euthyroid patients, with normal TSH, high free T4 and T3, and normal total T4 and T3 performed in a Roche Diagnostics ® COBAS 8000 device. When the test was repeated on a Siemens® Immulite device, the free and total hormones were within normal ranges. In the Roche Diagnostics ® assay, the presence of biotin or anti-Ruthenium or anti-streptavidin antibodies interferes with the formation of the complex responsible for the emission of light that allows inferring concentrations of thyroid hormones. The Siemens test works differently since the emission of light depends on the binding of T4 to an antibody conjugated with alkaline phosphatase not participating in the process biotin, streptavidin or ruthenium so this interference is avoided. This possible interference in immunoassays should be taken into account in case clinical manifestations differ from these laboratory determinations, to avoid a diagnosis and potential inappropriate treatment.


Resumen: La función tiroidea se evalúa midiendo tirotropina y concentraciones de hormonas tiroideas libres y totales. Existen interferencias con los resultados de inmunoensayos que pueden llevar a un diagnóstico incorrecto, de ellas, las más frecuentes son la unión de hormonas tiroideas a anticuerpos heterófilos, el factor reumatoide, anticuerpos anti Rutenio, la ingesta de biotina y anticuerpos anti estreptavidina. Se presentan tres casos de pacientes clínicamente eutiroideos, con TSH normal, T4 y T3 libres elevadas, y T4 y T3 totales normales realizadas en un equipo COBAS 8000 de Roche Diagnostics®. Cuando se repitió el ensayo en un equipo Immulite de Siemens®, las hormonas libres y totales estaban dentro de rangos normales. En el ensayo de Roche Diagnostics ®, la presencia de biotina o anticuerpos anti Rutenio o anti estreptavidina, interfiere con la formación del complejo responsable de la emisión de luz que permite inferir las concentraciones de las hormonas tiroideas. El ensayo de Siemens funciona de manera diferente ya que la emisión de luz depende de la unión de la T4 a un anticuerpo conjugado con fosfatasa alcalina no participando en el proceso biotina, estreptavidina o Rutenio por lo que se evita esta interferencia. Esta posible interferencia en inmunoensayos debe ser tenida en cuenta en caso de que las manifestaciones clínicas difieran de estas determinaciones de laboratorio, para evitar un diagnóstico y potencial tratamiento inadecuado.


Subject(s)
Humans , Female , Adult , Middle Aged , Thyroid Hormones/immunology , Thyroid Hormones/blood , Immunoassay/methods , Thyrotropin/immunology , Thyrotropin/blood , False Positive Reactions
4.
Rev. argent. endocrinol. metab ; 55(1): 43-56, mar. 2018. graf.
Article in Spanish | LILACS | ID: biblio-1248114

ABSTRACT

Esta revisión fue realizada con el fin de evaluar nuestros resultados de laboratorio así como aquellos de la literatura que constituyen, a nuestro entender, aportes significativos en el síndrome de ovarios poliquísticos (SOP). Nuestro especial énfasis será presentar las limitaciones de las metodologías empleadas por nuestro grupo, comparativamente a las reportadas por otros investigadores. La determinación de andrógenos, en particular de Testosterona (TT), es quizá la de mayor complejidad dado que los resultados con los diferentes inmunoensayos empleados en nuestro medio producen resultados muy variables por los diferentes métodos y aún entre laboratorios que usan la misma metodología. La técnica de referencia es la cromatografía líquida en tándem con espectrometría de masa (LC-MSMS), de difícil aplicación en laboratorios de análisis clínicos debido a su alto costo y la imposibilidad de resolver numerosas muestras. En estudios previos demostramos que de los métodos habitualmente usados para evaluar la TT circulante, solo en 2 inmunoensayos los resultados obtenidos fueron satisfactoriamente validados indirectamente según el criterio del Consenso de los Centros para el Control y Prevención de Enfermedades (CDC, USA) contra LC-MSMS, los cuales fueron comparables a dicha metodología con niveles superiores a 0,5 ng/ml. El SOP puede presentar factores de riesgo aumentados para la enfermedad cardiovascular y la diabetes II. Estos factores no están debidamente categorizados en función de los distintos fenotipos del SOP. Se evaluarán los principales analitos empleados con este objetivo y los nuevos que aporten elementos de mayor especificidad en este sentido


This review was performed in order to evaluate our laboratory results as well as those of the literature that constitute, in our opinion, significant contributions in these pathophysiologies. Our special emphasis will be on presenting the limitations of the methodologies used by our group, compared to those reported by other researchers. The determination of androgens, in particular Testosterone (TT), is perhaps the most complex since the results with the different immunoassays used in our environment produce very variable results by the different methods and even between laboratories that use the same methodology. The reference technique is LC-MSMS, difficult to apply in clinical analysis laboratories because of its high cost and the inability to solve numerous samples. In previous studies, we demonstrated that, in comparison to LC-MSMS with the usual methods for evaluating circulating TT, the results obtained in only 2 immunoassays were satisfactorily validated indirectly according to the criteria of CDC against LC-MSMS, which were comparable to that methodology with levels higher than 0.5 ng/ml. PCOS may have increased risk factors for cardiovascular disease and diabetes II. These factors are not properly categorized according to the different phenotypes of PCOS. The main analytes used for this purpose will be evaluated and new ones that contribute elements of greater specificity in this sense


Subject(s)
Humans , Female , Polycystic Ovary Syndrome/etiology , Polycystic Ovary Syndrome/physiopathology , Testosterone/analysis , Phenotype , Mass Spectrometry/methods , Immunoassay/methods , Chromatography, Liquid/methods
5.
Braz. j. med. biol. res ; 51(12): e7900, 2018. tab
Article in English | LILACS | ID: biblio-1039240

ABSTRACT

The impact of food restriction (FR) during 56 days on serum levels of cytokines in mice fed a high-fat diet (HFD) or high-carbohydrate diet (HCD) were evaluated. The amount of food was reduced 50% for HFD-FR and HCD-FR groups compared to mice receiving free access to HFD (HFD group) or HCD (HCD group). We quantified the serum levels of basic fibroblast growth factor, granulocyte-macrophage colony-stimulating factor, inducible protein 10, interferon γ, interleukin 1α (IL-1α), IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12, IL-13, IL-17, keratinocyte chemoattractant, macrophage inflammatory protein-1α, monocyte chemotactic protein 1, monokine induced by IFN-γ, and tumor necrosis factor α. Only IL-12 levels were lower (P<0.05), for both HFD-FR (HFD-FR vs HFD) and HCD-FR (HCD-FR vs HCD). Therefore, IL-12 levels could be considered a biological marker of the beneficial effects of FR.


Subject(s)
Animals , Rabbits , Interleukin-12/blood , Caloric Restriction/methods , Diet, High-Fat/methods , Food Deprivation/physiology , Diet, Carbohydrate Loading/methods , Animal Nutritional Physiological Phenomena/physiology , Reference Values , Time Factors , Body Weight , Immunoassay/methods , Biomarkers/blood , Cytokines/blood
6.
Arch. endocrinol. metab. (Online) ; 61(5): 460-463, Sept.-Oct. 2017. graf
Article in English | LILACS | ID: biblio-887594

ABSTRACT

ABSTRACT Measuring thyroid hormones is an important aspect for the study of metabolism and for monitoring diseases in both human and animal models. The traditional method for hormone measurement in rats is the radioimmunoassay (RIA). However, the RIA is associated with some practical disadvantages, including the use of radioactive material, the need for specialized equipment and expert staff, the short shelf-life of kits according to the half-life of the radioisotope and high costs. The objective of this study was to develop a new cost-effective method for measuring TSH levels in rats that avoids the use of radioactive material. We developed an in-house competitive immunoassay using a reference standard, polyclonal antibody produced in rabbits and biotinylated antigen. This method was tested in 64 Wistar rats that were divided into a control group (n = 41) and a group with hypothyroidism (n = 23). Our assay demonstrated an analytical sensitivity of 0.24 ng/mL (n = 12) and an intra-assay coefficient of variation (CV) of 8.9% for sera with TSH levels of 1.5 ng/mL and 13.2% for sera with TSH levels of 17.5 ng/mL (n = 14). The inter-assay CV was 13.5% for sera with TSH levels of 1.4 ng/mL and 14.5% for TSH levels of 18.2 ng/mL (n = 5). The analysis of mean TSH levels in control rats (5.06 ± 0.5701) and hypothyroid rats (51.09 ± 5.136) revealed a statistically significant difference (p < 0.001) between the groups. This method showed good sensitivity, can be automated and is low-cost compared with RIA. Our method offers a viable alternative for TSH measurement in rats.


Subject(s)
Animals , Male , Rabbits , Rats , Thyroid Diseases/diagnosis , Immunoassay/methods , Thyrotropin/blood , Thyroid Diseases/blood , Immunoassay/economics , Sensitivity and Specificity , Cost-Benefit Analysis , Rats, Wistar
7.
Arch. endocrinol. metab. (Online) ; 60(5): 420-425, Oct. 2016. tab, graf
Article in English | LILACS | ID: lil-798183

ABSTRACT

ABSTRACT Objective To evaluate the usefulness of a third-generation PTH assay in the diagnosis of primary hyperparathyroidism (PHPT). Subjects and methods Forty-one PHPT patients (4 men and 37 women) with 61.2 ± 10.9 (mean ± SD) years, were studied and had PTH levels measured with two different methods using the same immunochemiluminescent assay plataform (Elecsys 2010 System, Roche). We compared a second-generation assay (I-PTH) with a third-generation PTH assay (Bio-PTH). Two populations of 423 and 120 healthy adults with serum 25OHD levels above 25 ng/mL were used to define normal values in the I-PTH and Bio-PTH assays respectively. Results Normal PTH values based in the healthy adults population were 24.2-78.0 pg/mL for the I-PTH assay and 19.9-58.5 pg/mL for Bio-PTH assay. In PHPT patients, PTH values ranged from 67 to 553 pg/mL (median: 168 pg/mL) using the I-PTH assay and from 55 to 328 pg/mL (median: 111 pg/mL) using the Bio-PTH assay. Results obtained with the Bio-PTH assay were significantly lower (p < 0.0001, Wilcoxon). In general I-PTH and Bio-PTH showed highly significant correlation (r = 0.952, p < 0.0001). Passing–Bablok analysis gave a regression equation of Bio PTH = 13.44 + 0.59 x intact PTH. PHPT patients had 25OHD levels ranging from 4 to 36 ng/mL (mean 16.2 ng/mL); 35 subjects (85.3%) had values bellow 25 ng/mL. Conclusion Our results demonstrate that both second and third generation PTH methods are strongly correlated in PHPT patients and control subjects. Lower results with Bio-PTH tests are expected in function of the assay specificity determined by the amino-terminal antibody used.


Subject(s)
Humans , Male , Female , Middle Aged , Parathyroid Hormone/blood , Peptide Fragments/blood , Hyperparathyroidism, Primary/diagnosis , Hyperparathyroidism, Primary/blood , Reference Standards , Reference Values , Brazil , Immunoassay/methods , Biomarkers/blood , Case-Control Studies , Reproducibility of Results , Statistics, Nonparametric , Luminescent Measurements/methods
9.
Clinics ; 71(4): 226-231, Apr. 2016. tab, graf
Article in English | LILACS | ID: lil-781421

ABSTRACT

OBJECTIVES: To establish cut-off values for growth hormone concentrations using clonidine as a secretagogue and an immunochemiluminescent assay as the method of measurement and to analyze the response time as well as the influence of gender, nutritional status and pubertal stage. METHODS: A total of 225 tests were performed in 3 patient groups, categorized as group 1 (normal), group 2 (idiopathic short stature) and group 3 (growth hormone deficiency). Among the 199 disease-free individuals, 138 were prepubertal, and 61 were pubertal. Clonidine (0.1 mg/m2) was orally administered, and the growth hormone level was measured by immunochemiluminescent assay. The growth hormone peak and the difference between the growth hormone peak and the baseline level were then analyzed. Statistical analyses were performed using Student’s t-test or the Mann-Whitney test and Kruskal-Wallis test followed by Dunn’s post hoc test. Cut-off values were determined using a receiver operating characteristic curve. RESULTS: Group 1 and group 2 had no difference in growth hormone peak, gender, body mass index standard deviation score, or pubertal stage. Group 3 exhibited a significantly lower growth hormone peak than the other groups did. The receiver operating characteristic curve demonstrated that growth hormone concentrations ≥ 3.0 ng/mL defined responsiveness to clonidine. In total, 3.02% of individuals in group 1 and group 2 were considered false positive, i.e., these children lacked growth hormone deficiency and had a peak below 3.0 ng/mL. CONCLUSION: Clonidine-stimulated growth hormone concentrations ≥3 ng/mL, as measured by immunochemiluminescent assay, suggest responsiveness to the stimulus regardless of gender, body mass index standard deviation score or pubertal stage.


Subject(s)
Humans , Male , Female , Child, Preschool , Child , Adolescent , Adrenergic alpha-2 Receptor Agonists/pharmacology , Body Height , Clonidine/pharmacology , Growth Disorders/diagnosis , Growth Hormone/deficiency , Human Growth Hormone/blood , Case-Control Studies , Growth Disorders/blood , Growth Disorders/etiology , Growth Hormone/blood , Immunoassay/methods , Insulin-Like Growth Factor I/analysis , Luminescent Measurements/methods , Prospective Studies , ROC Curve
10.
Biomed Res Int ; 2015: 1-16, 2015. ilus
Article in English | LILACS, SES-SP, SESSP-IALPROD, SES-SP, SESSP-IALACERVO | ID: biblio-1022429

ABSTRACT

Schistosomiasis constitutes a major public health problem, with an estimated 200 million people infected worldwide. Many areas of Brazil show low endemicity of schistosomiasis, and the current standard parasitological techniques are not sufficiently sensitive to detect the low-level helminth infections common in areas of low endemicity (ALEs). This study compared the Kato-Katz (KK); Hoffman, Pons, and Janer (HH); enzyme-linked immunosorbent assay- (ELISA-) IgG and ELISA-IgM; indirect immunofluorescence technique (IFT-IgM); and qPCR techniques for schistosomiasis detection in serum and fecal samples, using the circumoval precipitin test (COPT) as reference. An epidemiological survey was conducted in a randomized sample of residents from five neighborhoods of Barra Mansa, RJ, with 610 fecal and 612 serum samples. ELISA-IgM (21.4%) showed the highest positivity and HH and KK techniques were the least sensitive (0.8%). All techniques except qPCR-serum showed high accuracy (82­95.5%), differed significantly from COPT in positivity , and showed poor agreement with COPT. Medium agreement was seen with ELISA-IgG (Kappa = 0.377) and IFA (Kappa = 0.347). Parasitological techniques showed much lower positivity rates than those by other techniques. We suggest the possibility of using a combination of laboratory tools for the diagnosis of schistosomiasis in ALEs.


Subject(s)
Schistosomiasis mansoni/diagnosis , Schistosomiasis mansoni/parasitology , Schistosomiasis mansoni/epidemiology , Aged, 80 and over , Brazil/epidemiology , Aged , Humans , Immunoassay/methods , Immunoassay/statistics & numerical data , Precipitin Tests/methods , Child , Child, Preschool , Population Surveillance/methods , Prevalence , Reproducibility of Results , Sensitivity and Specificity , Risk Assessment/methods , Adult , Clinical Laboratory Techniques/methods , Clinical Laboratory Techniques/statistics & numerical data , Endemic Diseases/statistics & numerical data , Young Adult , Infant , Middle Aged
11.
Braz. j. microbiol ; 45(4): 1357-1361, Oct.-Dec. 2014. ilus, tab
Article in English | LILACS | ID: lil-741287

ABSTRACT

Several reports showed outbreaks of histoplasmosis acquired while bat-inhabited caves were visited by tourists, miners or researchers. We evaluated the performance of double immunodifusion (DI) and immunoblotting (IB) assays, employed for the histoplasmosis outbreak elucidation occurred in Vale do Paraíba, São Paulo. The existence of epidemiologic link, four patients with clinical signs suggestive of histoplasmosis and mycological confirmation has made that all 35 individuals involved to the cave visit were subjected to serological evaluation. By DI, we observed reactivity against H. capsulatum antigen in a single serum examined nearly 20 days after exposure to fungal propagules. On the other hand, IB showed reactivity against H and M fractions in 50% of samples evaluated. The analysis of the second sample batch, collected two months after the exposure showed that 96.7% were reactive by DI with antibodies titers ranging from 1 to 16 and 100% of reactivity against H and M fractions, by IB, suggesting an acute infection. The analysis of the overall agreement between the methods showed to be reasonable (κ = 0.37). This study confirms the importance and efficacy of more sensitive methodologies, such as IB assay, to early elucidation of disease, especially in cases of patients without mycological information.


Subject(s)
Adolescent , Adult , Female , Humans , Male , Antibodies, Fungal/blood , Disease Outbreaks , Diagnostic Tests, Routine/methods , Histoplasma/immunology , Histoplasmosis/diagnosis , Histoplasmosis/epidemiology , Microbiological Techniques/methods , Brazil/epidemiology , Immunoassay/methods , Sensitivity and Specificity
12.
Rev. salud pública ; 16(6): 1-1, nov.-dez. 2014. ilus, tab
Article in Spanish | LILACS | ID: lil-745110

ABSTRACT

Objetivo Determinar los perfiles serológicos para el virus de hepatitis B, en donantes de sangre anti-HBc reactivo y antígeno de superficie no reactivo, provenientes de cuatro ciudades del país Métodos Se realizó un estudio prospectivo transversal, durante un período de 17 meses, aplicando el perfil serológico completo de la hepatitis B, en muestras de donantes con anti-HBc reactivo y antígeno de superficie de hepatitis B no reactivo. Los resultados fueron analizados utilizando Microsoft® Excel y Epiinfo V 3.5.1. Resultados Se encontró que el 75 % de los donantes reactivos para anti-HBc en los bancos de sangre, presentaban algún marcador adicional de exposición para el VHB; el 1,3 % de los donantes presentaban marcadores serológicos de infección crónica por hepatitis B y un caso que resultó reactivo solamente para antígeno de superficie de hepatitis B. Se halló perfil de vacunación en el 6,1 % de donantes, que fueron reactivos solamente para anticuerpo contra antígeno de superficie.Conclusiones. Se ratifica la importancia de la tamización de anti-HBc, a los donantes de sangre...(AU)


To assess the serological profiles for HBV in blood donors that were anti-HBc reactive and non-reactive to HBsAg in four Colombian cities. Materials and Methods A prospective transversal study was conducted during 17 months, applying a complete serological profile for HBV in samples from blood donors that were anti-HBc reactive and non-reactive to HBsAg, results were analyzed employing descriptive statistics using Microsoft Excel and Epiinfo V. 3.5.1. Results From donors reactive to anti-HBc, 75.0 % shown some additional infection marker for HBV. 1.3 % of blood donors had serological markers for chronical infection with hepatitis B, and a case had reactivity only for hepatitis B surface antigen (HBsAg). 6.1 % of donors showed a vaccination serological profile, only with reactivity to anti-HBsAg antibodies. Conclusions With this study, anti-HBc blood screening importance was confirmed...(AU)


Subject(s)
Humans , Blood Banks/methods , Blood Donors , Blood Transfusion/methods , Hepatitis B/diagnosis , Immunoassay/methods , Cross-Sectional Studies/instrumentation , Prospective Studies
13.
Arq. bras. med. vet. zootec ; 66(3): 713-720, 06/2014. tab, graf
Article in English | LILACS | ID: lil-718097

ABSTRACT

This study aimed to assess and standardize the ELISA and modified ToBI test in vitro methods in order to verify the potency of epsilon toxicoid in comparison with the in vivo TCP method. The following epsilon toxoids were used: NIBSC standard from batches 375/07, 532/08, 551/08, 373/07 and 378/07. These were evaluated using a TCP test, ELISA and ToBI tests. The results indicate that the correlation ratio between the dilutions of standard NIBSC toxicoid and absorbance values of 89.44% obtained with the ELISA method support the use of the curve to evaluate epsilon toxoids. However, it was observed that the absorbance values were similar for all toxoids, thus presenting no significant difference between higher and lower concentration toxoids. For the ToBI test, the correlation ratio of 96.76, obtained in the curve pattern, demonstrates the effectiveness of the curve to be used in the epsilon toxoid evaluation. The correlation ratio between the titration degrees of toxoids obtained through TCP and ToBI tests was higher than 90%. It is concluded that the type of ELISA test used does present discriminative power for toxoids with different concentrations, which does not support the use of this technique for such a purpose. The ToBI test can be used as a screening method for it is sensitive and effective to detect epsilon toxicoid produced by C. perfringens type D...


Teve-se por objetivo avaliar e padronizar as metodologias in vitro, ELISA e ToBI-test modificado, para a análise de toxoide épsilon, em comparação com a metodologia in vivo TCP. Foram utilizados os seguintes toxoides épsilon: padrão NIBSC e os lotes 375/07, 532/08, 551/08, 373/07 e 378/07, os quais foram avaliados por métodos in vivo, TCP, e in vitro, ELISA e ToBI-test. A análise do título de toxoide épsilon por meio dos métodos in vitro foi realizada a partir de uma curva-padrão, estabelecida previamente. Os principais resultados mostram que os valores de absorbância foram semelhantes para todos os toxoides, não apresentando diferença significativa entre os toxoides mais concentrados e menos concentrados. No ToBI-test, o coeficiente de correlação de 96,76%, obtido na curva-padrão, demonstra a eficiência da curva para avaliação do toxoide épsilon. O coeficiente de correlação entre os títulos de toxoide obtidos pelo TCP e ToBI-test foi superior a 90%. Conclui-se que o tipo de ELISA utilizado não apresenta poder discriminativo para toxoides com diferentes concentrações, inviabilizando a técnica para esse fim. O ToBI-test pode ser utilizado como um método de triagem sensível e eficaz para a detecção de toxoide épsilon de C. perfringens tipo D...


Subject(s)
Clostridium/isolation & purification , Enzyme-Linked Immunosorbent Assay , Toxoids/antagonists & inhibitors , Vaccines , Immunoassay/methods
14.
Rev. cuba. hig. epidemiol ; 51(1): 84-96, ene.-abr. 2013.
Article in Spanish | LILACS | ID: lil-671306

ABSTRACT

Entre las enfermedades relacionadas con el agua según su uso se encuentran las causadas por sustancias químicas y por agentes biológicos. Dentro de estas últimas, las ocasionadas por bacterias y protozoarios patógenos incrementan cada día la lista de enfermedades emergentes y reemergentes. Los métodos de ensayo para la determinación de microorganismos patógenos en el agua no han variado mucho en los últimos años, principalmente para los indicadores bacterianos de contaminación fecal, y por lo general se realizan por métodos convencionales. Sin embargo, existen situaciones, sobre todo en la aparición de brotes de enfermedades, en las que se hace necesario detectar el microorganismo patógeno en agua como posible agente causal, por lo que se ha recomendado el uso de métodos rápidos y confiables. Dentro de estos se encuentran los inmunoensayos, de los cuales los métodos por precipitación y aglutinación, los enzimoinmunoensayos, las técnicas de inmunofluorescencia directa e indirecta y la citometría de flujo son muy útiles en la detección de microorganismos en agua. Mención aparte merece la separación inmunomagnética o inmunocaptura como paso previo a otras técnicas avanzadas. Nos proponemos con este trabajo exponer las ventajas y desventajas de estos métodos, los principios en los cuales se basan y ejemplificar algunos de los más utilizados en microbiología de aguas, así como recalcar su importancia


Diseases related to the use of water may be caused by chemical substances or biological agents. Among the latter, a prominent role is played by pathogenic bacteria and protozoa, which constantly add to the list of emerging and re-emerging diseases. Assay methods to identify pathogenic microorganisms in water have not changed much in recent years, particularly with respect to bacterial indicators of fecal contamination, and tests are usually conducted by conventional methods. However, in certain situations, especially when a disease outbreak occurs, it is necessary to determine what pathogenic microorganism is the possible causal agent, and quick, reliable methods have been recommended to achieve this aim. These include immunoassays, among which precipitation and agglutination methods, enzyme immunoassays, direct and indirect immunofluorescence techniques and flow cytometry have proven very useful to detect microorganisms in water. Special mention should be made of immunomagnetic separation or immunocapture as a step preceding other advanced techniques. The present paper is aimed at presenting the advantages and disadvantages of these methods, as well as the principles on which they are based. Examples are provided of the methods most commonly used in water microbiology, highlighting their importance


Subject(s)
Bacteria/immunology , Water Quality/standards , Water Pollution/analysis , Eukaryota/immunology , Immunoassay/methods , Aquatic Microorganisms/adverse effects , Water Microbiology , Waterborne Diseases , Disease Outbreaks
15.
Lima; s.n; 2013. 43 p. ilus, tab.
Thesis in Spanish | LILACS, LIPECS | ID: lil-713899

ABSTRACT

Con el propósito de obtener una técnica altamente sensible y de bajo costo para el tamizaje de cisticercosis humana en zonas endémicas, se evaluó la prueba de Dot blot para la detección de anticuerpos en suero de personas infectadas por esta parasitosis. Para evaluar la eficiencia de la técnica, se emplearon sueros confirmados por Western blot, de los cuales, 60 procedían de pacientes con cisticercosis, 45 de pacientes con otras parasitosis y 20 de personas sanas, estos últimos para evaluar reacciones cruzadas. Se usó papel de nitrocelulosa, en el que el antígeno total de líquido vesicular de cisticercosis de Taenia solium fue fijado. A continuación, la tira fue incubada con el suero problema y luego con anti-IgG humano marcado con una enzima. El suero que tenia anticuerpos, al agregar un sustrato cromógeno, originó un producto insoluble que precipitó formando un punto en la zona donde se produjo la reacción antígeno-anticuerpo. La sensibilidad, especificidad, valor predictivo positivo y valor predictivo negativo de la prueba Dot blot fue de: 100 por ciento, 83 por ciento, 84 por ciento y 100 por ciento respectivamente. El índice de Kappa obtenido fue 0,98, lo que la califica como una prueba que tiene muy buena concordancia con el western blot. Por su alta sensibilidad, el uso de la prueba Dot blot parece ser un enfoque adicional para el descarte de cisticercosis, con la ventaja de la simplicidad y rapidez, susceptible de prepararse en regiones endémicas del país, constituyendo una buena alternativa como prueba de tamizaje y para estudios epidemiológicos, recomendándose por tanto su uso.


With the intention of obtaining a highly sensitive technology and of low cost for screening human cysticercosis in endemic zones, Dot blot test was evaluated for the detection of antibodies in persons' whey infected by this parasitosis. To evaluate the efficiency of the technique, there were used serums confirmed by Western blot, of which, 60 were coming from patients with cysticercosis, 45 of patients with other parasitosis and 20 of healthy persons, the above mentioned to evaluate crossed reactions. Nitrocellulose paper was used, in that the total antigen of liquid vesicular of cysticercosis of Taenia solium was fixed. Next, the strip was incubated with serum and then with anti-human IgG labeled with an enzyme. The serum had antibodies, by adding a suitable chromogenic substrate, resulted in an insoluble product which precipitated forming a point in the area where there was a reaction antigen-antibody. The sensibility, specificity, predictive positive value and predictive negative value of the test Dot blot was of: 100 per cent, 83 per cent, 84 per cent and 100 per cent respectively. The Kappa index obtained was 0.98, which qualifies it as a test that has very good conformity with the western blot. For his high sensibility, the use of the test Dot blot seems to be an additional approach for the discarded cards of cysticercosis with the advantage of the simplicity and rapidity, capable of to prepare in endemic regions of the country, constituting a good alternative as a screening test and for epidemiological studies, his use being recommended.


Subject(s)
Humans , Cysticercosis/diagnosis , Immunoassay/methods , Sensitivity and Specificity , Serum , Observational Study , Prospective Studies , Qualitative Research
16.
Yonsei Medical Journal ; : 1033-1039, 2013.
Article in English | WPRIM | ID: wpr-121783

ABSTRACT

PURPOSE: To investigate an association between the levels of serum thyroid-stimulating hormone (TSH)-receptor autoantibodies (TRAbs) and Graves' orbitopathy (GO) activity/severity scores, and compare the performance of three different TRAb assays in assessing the clinical manifestations of GO. MATERIALS AND METHODS: Cross-sectional study. Medical records of 155 patients diagnosed with GO between January 2008 and December 2010 were reviewed. GO activity was assessed by clinical activity score (CAS) and severity graded with the modified NOSPECS score by a single observer. Serum TRAb was measured by three different methods: 1st generation thyrotropin-binding inhibitor immunoglobulin (TBII) assay (TRAb1st); 3rd generation TBII assay (TRAb3rd); and biological quantitative assay of thyroid-stimulating immunoglobulin (TSI) using Mc4-CHO cells (Mc4-CHO TSI assay). Results were correlated with scores of activity/severity of thyroid eye disease. RESULTS: All three assays (TRAb1st, TRAb3rd, and Mc4-CHO TSI) yielded results that were significantly positively correlated with CAS (beta=0.21, 0.21, and 0.46, respectively; p<0.05) and proptosis (beta=0.38, 0.34, and 0.33, respectively; p<0.05). Mc4-CHO TSI bioassay results were significantly positively correlated with all GO severity indices (soft tissue involvement, proptosis, extraocular muscle involvement, and total eye score; beta=0.31, 0.33, 0.25, and 0.39, respectively; p<0.05). CONCLUSION: Mc4-CHO TSI bioassay was superior over the two TBIIs in assessing active inflammation and muscle restriction due to GO, whereas TBII assay would be sufficient for evaluation of patients with proptosis.


Subject(s)
Adult , Animals , Autoantibodies/blood , CHO Cells , Cricetulus , Cross-Sectional Studies , Female , Graves Ophthalmopathy/blood , Humans , Immunoassay/methods , Male , Middle Aged , Receptors, Thyrotropin/blood
17.
Article in English | WPRIM | ID: wpr-216012

ABSTRACT

Three automated immunoassay kits for anti-Hepatitis A Virus (HAV) IgM-Architect, (Abbott Laboratories, USA), Elecsys (Roche Diagnostics, Germany), and ADVIA Centaur (Siemens Healthcare Diagnostics Inc., USA)-were compared. We included 178 consecutive samples, for which an anti-HAV IgM test was requested at Seoul National University Hospital from September 2009 to January 2010. Reviewing of medical records, reverse transcription (RT)-PCR for HAV RNA, or total anti-HAV assay were performed on 16 (9.0%) samples with discrepant results. The percent agreements (kappas) of the Architect and ADVIA Centaur, Architect and Elecsys, and ADVIA Centaur and Elecsys kits were 96.6% (0.91), 96.6% (0.92), and 97.8% (0.94), respectively. Eight out of 16 discrepant samples showed gray-zone values in Architect but were nonreactive in the others. Slightly earlier seroconversion was suspected in Elecsys. The 3 assays showed comparable performances with excellent agreements in a tertiary care hospital setting.


Subject(s)
Hepatitis A virus/genetics , Hepatitis B/diagnosis , Hospitals, University , Humans , Immunoassay/methods , Immunoglobulin M/analysis , RNA, Viral/analysis , Reagent Kits, Diagnostic , Retrospective Studies , Reverse Transcriptase Polymerase Chain Reaction , Tertiary Healthcare
19.
Article in English | WPRIM | ID: wpr-180666

ABSTRACT

Non-human primate studies must be conducted prior to the clinical trial of xenotransplantation. In order to develop clinically applicable immune-modulatory regimen through non-human primate studies, close monitoring of xenogeneic immune responses is required. We adopted multiplex cytokine analysis in assessment of the immune status during the course of pig-to-non-human primate islet transplantation. This study aimed to assess the feasibility of this multiplex cytokine assay in the development of immune-modulatory regimen. Using this assay, we were able to detect different cytokines with a minimal usage of blood samples, and this allowed us to detect various immunological situations in the recipients. Detection of TNF-alpha surge (347.8 pg/mL) guided us to block TNF-alpha in the early phase of transplantation. Supportive information for in vivo efficacy of cytokine neutralizing antibody could be speculated by in vitro neutralization assay (1,250 pg/mL --> 0 pg/mL). In addition, periodic monitoring of cytokines in peripheral blood allowed the detection of the infection episode prior to other routine assays. These benefits of multiplex cytokine assay may be generally applied to other pre-clinical research, which is a prerequisite for clinical trials.


Subject(s)
Animals , Antibodies, Neutralizing/immunology , Blood Cell Count , Cytokines/blood , Immunoassay/methods , Interleukin-6/blood , Islets of Langerhans Transplantation/immunology , Macaca mulatta , Swine , Transplantation, Heterologous , Tumor Necrosis Factor-alpha/blood
20.
Article in English | WPRIM | ID: wpr-216689

ABSTRACT

A field applicable diagnostic technique, the dipstick assay, was evaluated for its sensitivity and specificity in diagnosing human Schistosoma mansoni infection. A monoclonal antibody (mAb) against S. mansoni adult worm tegumental antigen (AWTA) was employed in dipstick and sandwich ELISA for detection of circulating schistosome antigen (CSA) in both serum and urine samples. Based on clinical and parasitological examinations, 60 S. mansoni-infected patients, 30 patients infected with parasites other than schistosomiasis, and 30 uninfected healthy individuals were selected. The sensitivity and specificity of dipstick assay in urine samples were 86.7% and 90.0%, respectively, compared to 90.0% sensitivity and 91.7% specificity of sandwich ELISA. In serum samples, the sensitivity and specificity were 88.3% and 91.7% for dipstick assay vs. 91.7% and 95.0% for sandwich ELISA, respectively. The diagnostic efficacy of dipstick assay in urine and serum samples was 88.3% and 90.0%, while it was 90.8% and 93.3% for sandwich ELISA, respectively. The diagnostic indices of dipstick assay and ELISA either in serum or in urine were statistically comparable (P>0.05). In conclusion, the dipstick assay offers an alternative simple, rapid, non-invasive technique in detecting CSA or complement to stool examinations especially in field studies.


Subject(s)
Animals , Antibodies, Helminth , Antibodies, Monoclonal , Antigens, Helminth/blood , Diagnostic Tests, Routine/methods , Humans , Immunoassay/methods , Parasitology/methods , Point-of-Care Systems , Schistosoma mansoni/immunology , Schistosomiasis mansoni/diagnosis , Sensitivity and Specificity
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