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PublicouIrene Peralta Gama Alterado mais de 7 anos atrás
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Imunologia – 2ª parte Aplicações práticas (alguns exemplos)
Anticorpos Monoclonais Citometria de fluxo Com ou sem anticorpos monoclonais Métodos em Alergologia Basotest Determinação de IgE Alergograma CAST
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Anticorpos monoclonais
Anticorpos produzidos apenas por um clone de células Especificidade Reprodutibilidade
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Aplicações dos Ac monoclonais
Identificação e doseamentos Diagnóstico Humoral Fenotipagem celular / contagem Quantificação de proteínas em amostras Terapêutica Anti-neoplásica Anti-inflamatória Outras
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Populações linfocitárias
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Populações linfocitárias
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Populações linfocitárias
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TESTE DE ACTIVAÇÃO DE BASÓFILOS
marcador directo de activação celular CD 63 / CD 203c marcador indirecto de libertação de histamina Apresenta boa sensibilidade e especificidade B
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TESTES DE ACTIVAÇÃO DE BASÓFILOS CITOMETRIA DE FLUXO
Apreciação de reacções IgE mediadas Determinação da percentagem de basófilos activados Estimulação “in vitro” com alergénios específicos Boa correlação com outros testes “in vitro” e “in vivo” SPAIC-OUTUBRO /2003
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Testes Laboratoriais IgE
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IgE TOTAL _______________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________
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IgE TOTAL _______________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________
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UNICAP
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IgE Total IgE Específica Rast ImunoCAP Ac anti-IgE marcado IgE sérica
Fase sólida com Ac anti-IgE _______________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________ ImunoCAP
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Multi-Rast Phadiatop ImunoCAP
_______________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________ ImunoCAP
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Rabo de Gato Phleum Pratense Extracção
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Rabo de Gato Phleum Pratense Extracção
____________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________________ Padrão proteico de doentes alérgicos por immunobloting
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Allergograma (Phleum pratense)
Rabo de Gato Phleum Pratense Phl p1 Phl p5
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Alergénios recombinantes -técnica
poly-(A) Allergy-eliciting biological material Standard Allergen ImmunoCAP Extraction, Immobilization on solid phase mRNA extraction poly-(A) cDNA synthesis, cloning and selection of cDNA clones encoding individual allergenic proteins Recombinant Allergen Production of recombinant allergens, Immobilization on solid phase Standardization and research reagents Pharmacia-Diagnostics
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“MICROARRAY” DE ALERGÉNIOS: TÉCNICA
Clin Exp. Allergy 2003; 7-13 GAILL-2003
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DIAGNÓSTICO DA ALERGIA (Presente)
Valenta,R. et all, Clinical Experimental Allergy, 1999, vol 29, GAILL-2003
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ALERGÉNIOS RECOMBINANTES-DIAGNÓSTICO
Valenta,R. et all, Clinical Experimental Allergy, 1999, vol 29, GAILL-2003
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Cellular Antigen Stimulation Test
Whole-blood EDTA Dextran sedimentation and 90 minutes incubation Transfer leukocytes, centrifuge, remove/discard upper phase + stimulation buffer (IL-3) sLT sLT A A sLT A + Antigen A A A A A A Supernatants can be harvested, after centrifugation store at - 20 °C until ELISA test Leukocyte suspension, + antigen (40 min incubation)
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Cellular Antigen Stimulation Test
2. ELISA-Procedure Enzym sLT sLT sLT sLT Enzym sLT sLT sLT 2 h incubation, wash 3 times Supernatant + monoclonal antibody + substrate Enzym Color Reading at 405 nm sLT sLT sLT 30 min incubation, wash 3 times + stop-solution
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Cellular Antigen Stimulation Test (CAST)
Disponível para Antibióticos: Penicilina, Ampicilina /Amoxicilina Cefalosporina, Cefuroxime Trimethoprim, Sulfametoxazol Tetraciclina Ciprofloxacina Analgésicos: Aspirina, Diclofenac, Ibuprofeno, Indometacina Paracetamol, Metamizol Anestésicos Miorelaxantes
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Imunologia – 2ª parte Aplicações práticas (alguns exemplos)
Anticorpos Monoclonais Citometria de fluxo Com ou sem anticorpos monoclonais Métodos em Alergologia Basotest Determinação de IgE Alergograma CAST
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