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au:Dellavance, Alessandra

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Periostin as an important biomarker of inflammatory phenotype T2 in Brazilian asthma patients
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Baldo, Danielle Cristiane

; Romaldini, José Gustavo

; Pizzichini, Marcia Margaret Menezes

; Cançado, José Eduardo D.

; Dellavance, Alessandra

; Stirbulov, Roberto

Jornal Brasileiro de Pneumologia

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2023, Volume 49 N. 1 elocation e20220040

Abstract: En Pt | Text: En Pt | PDF: En | PDF: Pt

ABSTRACT Objective: The aim of this study was to assess the laboratory performance of periostin associated with a panel of biomarkers to identify the inflammatory phenotype of Brazilian asthma patients. Methods: We evaluated 103 Brazilian individuals, including 37 asthmatics and 66 nonasthmatic controls. Both groups underwent analyses for serum periostin, eosinophil levels in the peripheral blood, the fraction of exhaled nitric oxide (FeNO), total serum IgE, urinary leukotriene E4, and serum cytokines. Results: Higher levels of periostin (p = 0.005), blood eosinophils (p = 0.012), FeNO (p = 0.001), total IgE (p < 0.001), and IL-6 (p ≤ 0.001) were found in the asthmatic patients than the controls. Biomarker analyses by the ROC curve showed an AUC greater than 65%. Periostin (OR: 12,550; 95% CI: 2,498-63,063) and IL-6 (OR: 7,249; 95% CI: 1,737-30,262) revealed to be suitable asthma inflammation biomarkers. Blood eosinophils, FeNO, total IgE, IL-6, TNF, and IFN-g showed correlations with clinical severity characteristics in asthmatic patients. Periostin showed higher values in T2 asthma (p = 0.006) and TNF in non-T2 asthma (p = 0.029). Conclusion: The panel of biomarkers proposed for the identification of the inflammatory phenotype of asthmatic patients demonstrated good performance. Periostin proved to be an important biomarker for the identification of T2 asthma.

RESUMO Objetivo: O objetivo deste estudo foi de avaliar o desempenho laboratorial da periostina associada a um painel de biomarcadores para identificar o fenótipo inflamatório de pacientes brasileiros com asma. Métodos: Foram avaliados 103 indivíduos brasileiros, incluindo 37 asmáticos e 66 controles não asmáticos. Ambos os grupos foram submetidos a análises de periostina sérica, níveis de eosinófilos no sangue periférico, a fração exalada de óxido nítrico (FeNO), IgE sérica total, leucotrieno E4 urinário e citocinas séricas. Resultados: Maiores níveis de periostina (p = 0,005), eosinófilos periféricos (p = 0,012), FeNO (p = 0,001), IgE total (p < 0,001) e IL-6 (p ≤ 0,001) foram encontrados nos pacientes asmáticos do que nos controles. As análises de biomarcadores pela curva ROC mostraram uma AUC superior a 65%. A periostina (OR: 12.550; IC 95%: 2.498-63.063) e a IL-6 (OR: 7.249; IC 95%: 1.737-30.262) se mostraram biomarcadores adequados da inflamação da asma. Eosinófilos periféricos, FeNO, IgE total, IL-6, TNF e IFN-g apresentaram correlação com características clínicas de gravidade em pacientes asmáticos. A periostina teve valores mais elevados na asma T2 (p = 0,006) e o TNF na asma não T2 (p = 0,029). Conclusão: O painel de biomarcadores proposto para a identificação do fenótipo inflamatório de pacientes asmáticos demonstrou bom desempenho. A periostina provou ser um importante biomarcador para a identificação da asma T2.

https://doi.org/10.36416/1806-3756/e20220040

Anti-ribosomal P (anti-P) antibodies in patients with autoimmune hepatitis
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Gallo, Clarisse de Almeida

; Dellavance, Alessandra

; Gama, Raimundo Araújo

; Silva, Antônio Eduardo

; Silva, Ivonete Sandra de Souza e

; Andrade, Luis Eduardo Coelho

; Ferraz, Maria Lúcia Gomez

einstein (São Paulo)

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2023, Volume 21 elocation eAO0375

Abstract: En | Text: En | PDF: En

ABSTRACT Objective Published studies have shown associations between anti-ribosomal P (anti-P) antibody and systemic lupus erythematosus with hepatic manifestations. This has been reported also in autoimmune hepatitis. However, the consistency of the latter association remains controversial. This study aimed to evaluate the frequency of anti-P antibodies in autoimmune hepatitis using two different immunoassays. Methods One-hundred and seventy-seven patients with autoimmune hepatitis were screened, and 142 were analyzed for anti-P antibody positivity. The samples were first analyzed using two different immunoassays: enzyme-linked immunosorbent assay (ELISA) and chemiluminescence and then compared with a group of 60 patients with systemic lupus erythematous. The positive samples were subjected to western blot analysis. Results Anti-P was found in 5/142 autoimmune hepatitis cases (3.5%) by chemiluminescence and in none by ELISA. Among the five chemiluminescence-positive autoimmune hepatitis samples, on anti-P western blot analysis one was negative, two were weakly positive, and two were positive. In contrast, anti-P was detected in 10/60 patients with systemic lupus erythematosus (16.7%) and presented higher chemiluminescence units than the autoimmune hepatitis samples. Conclusion A low frequency of anti-P antibodies was observed in autoimmune hepatitis, suggesting that this test is not useful for the diagnosis or management of this disease.

https://doi.org/10.31744/einstein_journal/2023ao0375

VI Brazilian consensus guidelines for detection of anti-cell autoantibodies on HEp-2 cells
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Cruvinel, Wilson de Melo

; Andrade, Luis Eduardo Coelho

; Dellavance, Alessandra

; Ximenes, Antônio Carlos

; Bichara, Carlos David Araújo

; Mangueira, Cristóvão Luis Pitangueira

; Bonfá, Eloísa

; Brito, Fabiano de Almeida

; Mariz, Henrique Ataíde

; Anjos, Lisiane Maria Enriconi dos

; Pasoto, Sandra Gofinet

; Valim, Valeria

; Santos, Wilton Ferreira Silva dos

; Gomes, Clayson Moura

; Neves, Roberpaulo Anacleto

; Francescantonio, Paulo Luiz Carvalho

Advances in Rheumatology

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2022, Volume 62 elocation 34

Abstract: En | Text: En | PDF: En

Abstract Background: The VI Brazilian Consensus on Autoantibodies against HEp-2 cells for determination of autoantibodies against cellular constituents on HEp-2 cells was held on September, 2019, in Fortaleza (CE, Brazil). The guidelines in this edition were formulated by the group of Brazilian experts discussing the classification of complex patterns, the classification of the nuclear discrete dots (few and multiple), the identification of the discrete fine speckled pattern (AC-4a) and improvements on the ANA report. Mainbody: Sixteen Brazilian researchers and experts from universities and clinical laboratories representing the various geographical regions of Brazil participated in the meeting. Four main topics were discussed: (1) How to classify patterns with fluorescence in more than one cell compartment considering three relevant categoris: composite patterns, mixed patterns and multiple patterns; (2) The splitting of the discrete nuclear dots pattern into the multiple discrete nuclear dots (AC-6) and few discrete nuclear dots (AC-7) patterns, respectively; (3) Inclusion of a novel nuclear pattern characterized by discrete fine speckled pattern highly associated with antibodies to SS-A/Ro60, classified as AC-4a. In addition, adjustments on the Brazilian Consensus nomenclature were implemented aiming to harmonize the designation of some patterns with the International Consensus on ANA Patterns (ICAP). Furthermore, the designations of the PCNA-like pattern (AC-13), CENP-F-like pattern (AC-14) and Topo I-like pattern (AC-29) were adjusted in accordance to ICAP. Finally, there was a recommendation for adjustment in the test report in order to address the status of nuclear envelope staining. For all topics, the aim was to establish specific guidelines for laboratories and clinicians. All recommendations were based on consensus among participants. All recommendations from the V Consensus were maintained and there was relevant progress in the BCA/HEp-2 guidelines and further harmonization with ICAP. Conclusion: The VI BCA/HEp-2 edition was successful in establishing important recommendations regarding the classification of complex patterns, in supporting the identification of a novel pattern within the AC-4 group and in the harmonization process with the ICAP terminology.

https://doi.org/10.1186/s42358-022-00266-z

Antineutrophil cytoplasmic antibody profiles differ according to type of primary sclerosing cholangitis and autoimmune hepatitis
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Crescente, Juliana Goldbaum

; Dellavance, Alessandra

; Diniz, Marcio Augusto

; Carrilho, Flair Jose

; Andrade, Luis Eduardo Coelho de

; Cançado, Eduardo Luiz Rachid

Clinics

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2021, Volume 76 elocation e2228

Abstract: En | Text: En | PDF: En

OBJECTIVES: To determine the frequency of the antineutrophil cytoplasmic antibodies (ANCA), antiproteinase-3 and antimyeloperoxidase, in primary sclerosing cholangitis (PSC) with or without inflammatory bowel disease (IBD+ or IBD-) and in different types of autoimmune hepatitis (AIH). Additionally, to verify the agreement between ANCA patterns by indirect immunofluorescence and their antigenic specificities by ELISA. METHODS: For this study, 249 patients were enrolled (42 PSC/IBD+; 33 PSC/IBD-; 31 AIH type-1; 30 AIH type-2; 31 AIH type-3; 52 primary biliary cirrhosis; 30 healthy controls) whose serum samples were tested for ANCA autoantibodies. RESULTS: There were fewer female subjects in the PSC/IBD- group (p=0.034). Atypical perinuclear-ANCA was detected more frequently in PSC/IBD+ patients than in PSC/IBD- patients (p=0.005), and was significantly more frequent in type-1 (p<0.001) and type-3 AIH (p=0.012) than in type-2 AIH. Proteinase-3-ANCA was detected in 25 samples (only one with cytoplasmic-ANCA pattern), and more frequently in PSC/IBD+ than in PSC/IBD- patients (p=0.025). Myeloperoxidase-ANCA was identified in eight samples (none with the perinuclear-ANCA pattern). Among the 62 reactive samples for atypical perinuclear-ANCA, 13 had antigenic specific reactions for proteinase-3 and myeloperoxidase. CONCLUSIONS: PSC/IBD+ differed from PSC/IBD- in terms of sex and proteinase 3-ANCA and atypical perinuclear-ANCA reactivity, the latter of which was more frequently detected in type-1 and type-3 AIH than in type-2 AIH. There was no agreement between ANCA patterns and antigenic specificities in IBD and autoimmune liver diseases, which reinforces the need for proteinase-3 and myeloperoxidase antibody testing.

https://doi.org/10.6061/clinics/2021/e2228 431 downloads

V Brazilian consensus guidelines for detection of anti-cell autoantibodies on hep-2 cells
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Cruvinel, Wilson de Melo

; Andrade, Luis Eduardo Coelho

; von Mühlen, Carlos Alberto

; Dellavance, Alessandra

; Ximenes, Antônio Carlos

; Bichara, Carlos David

; Bueno, Cleonice

; Mangueira, Cristóvão Luis Pitangueira

; Bonfá, Eloísa

; Brito, Fabiano de Almeida

; Flumian, Fernanda Bull

; Silva, Glaucielen Gomes da

; Rêgo, Jozelia

; Anjos, Lisiane Maria Ericoni dos

; Slhessarenko, Natasha

; Pasoto, Sandra Gofinet

; Neves, Suzane Pretti Figueiredo

; Valim, Valéria

; Santos, Wilton Silva dos

; Francescantonio, Paulo Luiz Carvalho

Advances in Rheumatology

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2019, Volume 59 elocation 28

Abstract: En | Text: En | PDF: En

Abstract Background: The V Brazilian Consensus for determination of autoantibodies against cellular constituents on HEp-2 cells, held in Brasilia (DF, Brazil) on August 27, 2016, discussed the harmonization between the Brazilian Consensus on ANA (BCA) guidelines and the International Consensus on ANA Patterns (ICAP) recommendations (www.anapatterns.org). Initial guidelines were formulated by the group of Brazilian experts with the purpose of guiding and enabling Brazilian clinical laboratories to adopt recommendations and to provide a common standard for national and international consensuses. Mainbody: Twenty Brazilian researchers and experts from universities and clinical laboratories representing the various geographical regions of the country participated in the meeting. Three main topics were discussed, namely the harmonization between the BCA guidelines and latest recommendations of the ICAP initiative, the adjustment of the terminology and report on HEp-2 patterns, and a reassessment of quality assurance parameters. For the three topics, our aim was to establish specific guidelines. All recommendations were based on consensus among participants. There was concrete progress in the adjustment of the BCA guidelines to match the ICAP guidelines. To a certain extent, this derives from the fact that ICAP recommendations were largely based on the algorithm and recommendations of the IV Brazilian ANA Consensus, as consistently recognized in the ICAP publications and presentations. However, although there is great overlap between the two Consensuses, there are some point divergences. These specific items were individually and extensively discussed, and it was acknowledged that in several points ICAP improved recommendations previously issued by the Brazilian ANA Consensus and these changes were readily implemented. Regarding some specific topics, the BCA panel of experts felt that the previously issued recommendations remained relevant and possibly will require further discussion with ICAP. The term anti-cell antibodies was adopted as the recommended designation, recognizing that the assay addresses antibodies against antigens in the nucleus and in other cell compartments. However, the acronym ANA HEp-2 was maintained due to historical and regulatory reasons. It was also signalized that the latest trend in ICAP is to adopt the term Indirect Immunofluorescent Assay on HEp-2 cell substrate (HEp-2 IIFA). In addition, the quality assurance strategies previously presented were ratified and emphasized. Conclusion: The V BCA edition was successful in establishing an overall harmonization with the ICAP recommendations for interpretation of the HEp-2 IIFA test, pinpointing the perspectives in filling the remaining gaps between both initiatives.

https://doi.org/10.1186/s42358-019-0069-5 1618 downloads

Implementation of the ANA HEp-2 consensus guidelines in Brazilian clinical laboratories
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Silva, Glaucielen G.

; Gomes, Clayson M.

; Dellavance, Alessandra

; Francescantonio, Paulo Luiz C.

; Andrade, Luís Eduardo C.

; Cruvinel, Wilson M.

Jornal Brasileiro de Patologia e Medicina Laboratorial

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nov 2017, Volume 53 N. 6 Pages 368 - 376

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

RESUMO Introdução: A pesquisa de autoanticorpos em células HEp-2 representa uma relevante ferramenta no auxílio diagnóstico de doenças autoimunes, especialmente as reumáticas. Em virtude dos avanços metodológicos, a técnica aumentou gradativamente a sensibilidade, bem como a necessidade de padronização. Objetivo: Avaliar a implantação das recomendações dos consensos brasileiros de pesquisa de autoanticorpos em células HEp-2. Métodos: Preenchimento de formulário em plataforma virtual direcionada aos laboratórios clínicos que realizam a metodologia. Os participantes responderam a um questionário sobre a adoção das diretrizes dos consensos brasileiros, detalhando os aspectos técnicos, o controle de qualidade, a leitura de lâminas e a emissão de laudos. Resultados: Participaram do estudo 53 laboratórios responsáveis por mais de 300 mil testes de fator antinuclear (FAN)/mês; mais da metade (58,5%) informou adotar integralmente as recomendações dos consensos. A maioria (83,1%) utiliza a diluição 1:80 para triagem, e 75,5% dos laboratórios, programas de educação e controle de qualidade. Apenas 39,6% utilizam mais de uma marca de kit para a realização do teste, e 32,1% não relataram observar todas as fases do ciclo celular na leitura da lâmina. O estudo detectou ainda discreta heterogeneidade entre participantes na identificação de padrões. Conclusão: Os resultados evidenciam a adoção das recomendações dos consensos de forma absoluta pela maioria dos laboratórios participantes, bem como a necessidade de aperfeiçoamento em alguns aspectos relevantes para a qualidade do ensaio.

ABSTRACT Introduction: The detection of autoantibodies in HEp-2 cells represents a relevant tool for the diagnosis of autoimmune diseases, especially rheumatic autoimmune diseases. As a result of the methodological advances, the technique gradually increased the sensitivity, as well as the need for standardization. Objective: To evaluate the implementation of the Brazilian Consensus recommendations for autoantibody determination in HEp-2 cells. Methods: A structured form in a virtual platform was filled in by experts in clinical laboratories that carry out the methodology across the country. The questionnaire addressed the adoption of the Brazilian Consensus guidelines, detailing the technical aspects, quality control, the strategy for reading slides and the release of reports. Results: The study included 53 laboratories responsible for more than 300,000 antinuclear antibody (ANA) tests/month; more than half (58.5%) reported fully adopting the recommendations of the Brazilian Consensus. The majority (83.1%) used the 1:80 for screening dilution, and 75.5% of laboratories, perform education and quality control programs. Only 39.6% reported using more than one kit brand to perform the test, and 32.1% did not report observing all phases of the cell cycle during slide reading. The study also detected some heterogeneity among participants in the identification of patterns. Conclusion: The results confirm the adoption of the Brazilian Consensus recommendations by most of participating laboratories, although with variable extent. There is need for improvement in some aspects, especially those related to the quality control.

https://doi.org/10.5935/1676-2444.20170059 1313 downloads

IV Consenso Brasileiro para pesquisa de autoanticorpos em células HEp-2
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Francescantonio, Paulo Luiz Carvalho

; Cruvinel, Wilson de Melo

; Dellavance, Alessandra

; Andrade, Luis Eduardo Coelho

; Taliberti, Ben Hur

; von Mühlen, Carlos Alberto

; Bichara, Carlos David Araújo

; Bueno, Cleonice

; Mangueira, Cristóvão Luis Pitangueira

; Carvalho, Darlene Gonçalves

; Bonfá, Eloísa S.D. de O.

; Brito, Fabiano de Almeida

; Araújo, Flávia Ikeda e

; Rêgo, Jozelia

; Pereira, Kaline Medeiros Costa

; Anjos, Lisiane Maria Enriconi dos

; Bissoli, Maria de Fatima

; Santiago, Mittermayer Barreto

; Maluf, Natalya Zaidan

; Alvarenga, Rossana Rassi

; Neves, Suzane Pretti Figueiredo

; Valim, Valeria

; Santos, Wilton Silva dos

Revista Brasileira de Reumatologia

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feb 2014, Volume 54 N. 1 Pages 44 - 50

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

Objetivo: O IV Consenso Brasileiro para Pesquisa de Autoanticorpos em Células HEp-2 (FAN) realizado em Vitória (ES), no dia 18 de setembro de 2012, objetivou discutir estratégias e recomendações relacionadas ao procedimento técnico, à padronização e à interpretação dos resultados da pesquisa de autoanticorpos em células HEp-2. Métodos: Participaram do evento 23 pesquisadores e especialistas de Universidades e laboratórios brasileiros. Foram abordados diferentes tópicos, discutidos amplamente a fim de se estabelecer recomendações específicas. Resultados e conclusão: O IV Consenso integrou à árvore de decisão o padrão citoplasmático em Anéis e Bastões, o padrão nuclear pontilhado Quasi-homogêneo (QH) e o padrão misto CENP-F. Discutiu-se ainda a necessidade de atenção para a classificação do padrão misto relacionado à presença de anticorpos anti-DNA topoisomerase I (Scl70), compreendendo os componentes nuclear pontilhado fino, nucleolar homogêneo, NOR na placa metafásica e citoplasmático pontilhado fino. Foram sugeridas diretrizes para o controle de qualidade do teste, diluição de triagem e diluição de esgotamento, e foi emitido alerta quanto à necessidade de atenção em relação à heterogeneidade de substratos disponíveis no mercado e a utilização de metodologias automatizadas para detecção de autoanticorpos.

Objective: The Fourth Brazilian Consensus for Autoantibodies Screening in HEp-2 Cells (ANA) was held in Vitória, Espírito Santo, and aimed to discuss strategies and recommendations about the technique, standardization, interpretation and quality control of the indirect immunofluorescence reaction on HEp-2 cells. Methods: Twenty three ANA experts from university centers and private laboratories in different areas from Brazil discussed and agreed upon recommendations for the fourth edition of the Brazilian Consensus for Autoantibodies Screening in HEp-2 Cells. Results and conclusion: The 4th ANA Consensus included three novel patterns into the existing algorithm (cytoplasmic Rods and Rings, nuclear Quasi-homogeneous, and CENP-F). Emphasis was given to the need of attention in describing the peculiar mixed pattern elicited by anti-DNA topoisomerase I (Scl-70) autoantibodies, comprising nuclear fine specked, nucleolar homogeneous pattern, NOR staining in metaphase plates, and cytoplasmic fine speckled patterns. The group also emphasized the need for continuous quality control in indirect immunofluorescence assays, the establishment of screening dilutions, as well as conjugate titration. An alert was made regarding the heterogeneity of commercial kits in defining patterns and the use of solid phase methodologies to determine the presence of autoantibodies.

https://doi.org/10.1016/j.rbr.2013.10.001 28365 downloads

Variability in the recognition of distinctive immunofluorescence patterns in different brands of HEp-2 cell slides
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Dellavance, Alessandra

; Cruvinel, Wilson de Melo

; Francescantonio, Paulo Luiz Carvalho

; Mangueira, Cristovão Luis Pitangueira

; Drugowick, Inês Cristina

; Rodrigues, Silvia Helena

; Andrade, Luis Eduardo Coelho

Jornal Brasileiro de Patologia e Medicina Laboratorial

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jun 2013, Volume 49 N. 3 Pages 182 - 190

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

INTRODUÇÃO: A imunofluorescência indireta (IFI) utilizando células HEp-2 como substrato antigênico é o teste padrão-ouro para a pesquisa de autoanticorpos contra antígenos celulares. Contudo, as condições de cultivo, fixação e permeabilização celular interferem diretamente na preservação e na distribuição espacial dos antígenos. Portanto, pode-se presumir que distintas condições no preparo das células possam interferir no reconhecimento dos padrões de imunofluorescência associados aos diversos autoanticorpos. OBJETIVO: Avaliar um painel de amostras de soro representativo de padrões nuclear, nucleolar, citoplasmático, de aparelho mitótico e de placa cromossômica em substratos de células HEp-2 de diferentes fornecedores. MATERIAIS E MÉTODOS: Sete observadores blindados e independentes de três centros de referência avaliaram 17 amostras que apresentavam diferentes padrões nucleares, nucleolares, citoplasmáticos e associados ao aparelho mitótico em lâminas com células HEp-2 de oito procedências. As lâminas foram codificadas para manter a confidencialidade das marcas, bem como dos centros participantes. RESULTADOS: Os 17 padrões de imunofluorescência em células HEp-2 foram reconhecidos na maioria dos substratos. No entanto, alguns substratos mostraram déficit na apresentação de alguns padrões (nuclear pontilhado grosso/U1-ribonucleoprotein associado a anticorpos contra o RNP (U1 ribonucleoproteína), sugestivo da presença de anticorpos anti-CENP-F (proteína centromérica F), sugestivo de anticorpos contra antígenos de célula em proliferação (proliferating cell nuclear antigen [PCNA]), citoplasmático pontilhado fino associado a anticorpos anti-Jo-1 (histidil sintetase), anti-NuMA-1 (nuclear mitotic apparatus protein 1) e anti-NuMA-2 (nuclear mitotic apparatus protein 2). CONCLUSÃO: Em que pese a boa qualidade geral dos substratos avaliados, existe divergência nos resultados obtidos entre os diferentes substratos comerciais. As variações observadas podem ser devidas aos lotes avaliados, portanto não se pode generalizar para as respectivas marcas. Recomenda-se que cada novo lote ou marca de lâmina sejam testados com diferentes soros referência representativos dos diversos padrões.

INTRODUCTION: Indirect immunofluorescence on HEp-2 cells is considered the gold standard for the detection of autoantibodies against cellular antigens. However, the culture conditions, cell fixation and permeabilization processes interfere directly in the preservation and spatial distribution of antigens. Therefore, one can assume that certain peculiarities in the processing of cellular substrate may affect the recognition of indirect immunofluorescence patterns associated with several autoantibodies. OBJECTIVE: To evaluate a panel of serum samples representing nuclear, nucleolar, cytoplasmic, mitotic apparatus, and chromosome plate patterns on HEp-2 cell substrates from different suppliers. MATERIALS AND METHODS: Seven blinded observers, independent from the three selected reference centers, evaluated 17 samples yielding different nuclear, nucleolar, cytoplasmic and mitotic apparatus patterns on HEp-2 cell slides from eight different brands. The slides were coded to maintain confidentiality of both brands and participating centers. RESULTS: The 17 HEp-2 cell patterns were identified on most substrates. Nonetheless, some slides showed deficit in the expression of several patterns: nuclear coarse speckled/U1-ribonucleoprotein associated with antibodies against RNP (U1RNP), centromeric protein F (CENP-F), proliferating cell nuclear antigen (PCNA), cytoplasmic fine speckled associated with anti-Jo-1 antibodies (histidyl synthetase), nuclear mitotic apparatus protein 1 (NuMA-1) and nuclear mitotic apparatus protein 2 (NuMA-2). CONCLUSION: Despite the overall good quality of the assessed HEp-2 substrates, there was considerable inconsistency in results among different commercial substrates. The variations may be due to the evaluated batches, hence generalizations cannot be made as to the respective brands. It is recommended that each new batch or new brand be tested with a panel of reference sera representing the various patterns.

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III Consenso Brasileiro para Pesquisa de Autoanticorpos em Células HEp-2: perspectiva histórica, controle de qualidade e associações clínicas
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Francescantonio, Paulo Luiz Carvalho

; Andrade, Luis Eduardo Coelho

; Cruvinel, Wilson de Melo

; Araújo, Flávia Ikeda e

; Dellavance, Alessandra

; Gabriel Júnior, Alexandre

; Nuccitelli, Barbara

; Taliberti, Ben Hur

; von Mühlen, Carlos Alberto

; Bichara, Carlos David Araújo

; Santos, Cláudio Henrique Ramos dos

; Bueno, Cleonice

; Yano, Cristiane Martinez

; Mangueira, Cristóvão Luis Pitangueira

; Carvalho, Darlene Gonçalves

; Cardoso, Elizângela

; Bonfá, Eloísa

; Rassi, Gustavo Gabriel

; Mundim, Hugo Mendonça

; Bendet, Izidro

; Rego, Jozélia

; Vieira, Lisiane Maria Enriconi dos Anjos

; Barbosa, Maria Ordália Ferro

; Sugiyama, Mitiko

; Santiago, Mittermayer Barreto

; Slhessarenko, Natasha

; Silva, Nilzio Antônio da

; Jarach, Renata

; Suda, Roberto

; Levy, Roger Abramino

; Sampaio, Silvia Oliveira

; Neves, Suzane Pretti Figueiredo

; Santos, Wilton Silva dos

; Nóbrega, Yanna K. de M.

Jornal Brasileiro de Patologia e Medicina Laboratorial

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jun 2009, Volume 45 N. 3 Pages 185 - 199

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

OBJETIVO: O III Consenso Brasileiro para Pesquisa de Autoanticorpos em Células HEp-2 (FAN) objetivou discutir estratégias para controlar a qualidade do ensaio, promover a atualização das associações clínicas dos diversos padrões e avaliar as dificuldades de implantação do II Consenso ocorrido no ano de 2002. MÉTODOS: Nos dias 13 e 14 de abril de 2007 participaram do encontro em Goiânia pesquisadores e especialistas de diversos centros universitários e laboratórios clínicos de diferentes regiões do Brasil, com o propósito de discutir e aprovar as recomendações que visam a melhores padronização, interpretação e utilização do ensaio pelos clínicos. Foram convidados como ouvintes representantes comerciais de diferentes empresas produtoras de insumos para realização do teste de FAN. RESULTADOS E CONCLUSÃO: Dada a heterogeneidade de microscópios e reagentes disponíveis no mercado, o III Consenso enfatizou a necessidade do controle de qualidade em ensaios de imunofluorescência indireta. Foram também feitas algumas adequações na terminologia utilizada para classificar os diferentes padrões. Finalmente, foi realizada uma atualização das associações clínicas com finalidade de facilitar cada vez mais o melhor uso do ensaio pelos clínicos.

OBJECTIVE: The Third Brazilian Consensus for Autoantibodies Screening in HEp-2 Cells (ANA) had as purpose the evaluation of difficulties in the accomplishment of the 2nd Consensus recommendations that took place in the year of 2002, the discussion of strategies for quality control of the assay and the discussion of an update of the clinical associations of the several immunofluorescent patterns. METHODS: Several ANA experts from university centers and private laboratories in different areas in Brazil joined the workshop in Goiânia on 2007 April 13 and 14 with the purpose of discussing and approving the recommendations for standardization, interpretation and use of the test by physicians. Commercial representatives of different ANA slide brands were also invited as listeners to the workshop. RESULTS AND CONCLUSION: The 3rd ANA Consensus emphasized the need for quality control in indirect immunofluorescent assays since there is a considerable heterogeneity of available microscopes and reagents. It also promoted adaptations in the previously approved terminology used to classify the different patterns and finally updated the clinical associations of the several patterns with the purpose of providing guidance for interpretation of the assay by clinical pathologists and assistant physicians.

21533 downloads

10.

3º Consenso Brasileiro para pesquisa de autoanticorpos em células HEp-2 (FAN): recomendações para padronização do ensaio de pesquisa de autoanticorpos em células HEp-2, controle de qualidade e associações clínicas
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Dellavance, Alessandra

; Gabriel Júnior, Alexandre

; Nuccitelli, Barbara

; Taliberti, Ben Hur

; von Mühlen, Carlos Alberto

; Bichara, Carlos David Araújo

; Santos, Cláudio Henrique Ramos dos

; Bueno, Cleonice

; Yano, Cristiane Martinez

; Mangueira, Cristóvão Luis Pitangueira

; Carvalho, Darlene Gonçalves

; Cardoso, Elizângela

; Bonfá, Eloísa

; Araújo, Flávia Ikeda e

; Rassi, Gustavo Gabriel

; Mundim, Hugo Mendonça

; Bendet, Izidro

; Rego, Jozélia

; Vieira, Lisiane Maria Enriconi dos Anjos

; Andrade, Luis Eduardo Coelho

; Barbosa, Maria Ordália Ferro

; Sugiyama, Mitiko

; Santiago, Mittermayer Barreto

; Slhessarenko, Natasha

; Silva, Nilzio Antônio da

; Francescantonio, Paulo Luiz Carvalho

; Jarach, Renata

; Suda, Roberto

; Levy, Roger Abramino

; Sampaio, Silvia Oliveira

; Neves, Suzane Pretti Figueiredo

; Cruvinel, Wilson de Melo

; Santos, Wilton Silva dos

; Nóbrega, Yanna K. de M.

Revista Brasileira de Reumatologia

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apr 2009, Volume 49 N. 2 Pages 89 - 98

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

OBJETIVO:O 3º Consenso Brasileiro para pesquisa de autoanticorpos em Células HEp-2 (FAN) teve como propósito avaliar as dificuldades de implantação do 2º Consenso ocorrido no ano de 2002, discutir estratégias para controlar a qualidade do ensaio e promover a atualização das associações clínicas dos diversos padrões. MÉTODOS:Participaram do encontro em Goiânia nos dias 13 e 14 de abril de 2008 pesquisadores e especialistas de diversos centros universitários e laboratórios clínicos de diferentes regiões do Brasil, com o propósito de discutir e aprovar as recomendações que visam à melhor padronização, interpretação e utilização do ensaio pelos clínicos. Representantes comerciais de diferentes empresas produtoras de insumos para realização do teste de FAN foram convidados como ouvintes. RESULTADOS E CONCLUSÕES: O 3º Consenso enfatizou a necessidade do controle de qualidade em imunofluorescência dada a heterogeneidade de microscópios e reagentes disponíveis no mercado, promoveu adequações na terminologia utilizada para classificar os diferentes padrões e, finalmente, atualizou as associações clínicas com finalidade de facilitar cada vez mais o melhor uso do ensaio pelos clínicos.

OBJECTIVE: The Third Brazilian Consensus for autoantibodies Screening in HEp-2 cells had as purpose the evaluation of difficulties in the accomplishment of the 2nd Consensus recommendations that took place in the year of 2002, the discussion of strategies for quality control of the assay and the promotion of an update of the clinical associations of the several immunofluorescent patterns. METHODS:Several ANA experts from university centers and private laboratories in different areas in Brazil joined the workshop in Goiânia on 2008 April 13 and 14 with the purpose of discussing and approving the recommendations for standardization, interpretation and use of the test by physicians. Commercial representatives of different ANA slide brands were also invited as listeners to the workshop. RESULTS AND CONCLUSIONS: The 3rd Consensus emphasized the need for quality control in indirect immunofluorescent since there is a considerable heterogeneity of available microscopes and reagents. It also promoted adaptations in the previously approved terminology used to classify the different patterns and finally updated the clinical associations of the several patterns with the purpose of providing guidance for interpretation of the assay by clinical pathologists and assistant physicians.

63793 downloads

11.

Importância do padrão de fluorescência na interpretação do teste do FAN: o caso do padrão pontilhado fino denso
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Dellavance, Alessandra

; Leser, Paulo Guilherme

; Andrade, Luís Eduardo Coelho

Revista da Associação Médica Brasileira

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oct 2007, Volume 53 N. 5 Pages 439 - 445

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

A pesquisa de anticorpos contra antígenos celulares (FAN-HEp-2), historicamente conhecida como FAN (fator antinúcleo), tem passado por modificações metodológicas que exigem reflexão sobre suas características de sensibilidade e especificidade, para que haja uma adequada valorização dos resultados obtidos. Atualmente, o exame é marcado por elevada sensibilidade e questionável especificidade, resultando em elevado número de exames positivos em indivíduos não auto-imunes. O reconhecimento de que uma fração substancial da população apresenta o teste de FAN-HEp-2 reagente ocasiona a necessidade de busca de características que possam distinguir os testes positivos em pacientes com doença auto-imune daqueles observados em indivíduos não auto-imunes. Entre essas características, encontram-se o título da reação e o padrão de fluorescência observado. Em particular, deve-se ter em mente que o padrão de fluorescência é resultante do reconhecimento de diferentes estruturas celulares pelos anticorpos e, portanto, o padrão de fluorescência fornece uma indicação preliminar dos possíveis antígenos que estão sendo reconhecidos. Um exemplo emblemático é representado pelo padrão nuclear pontilhado fino denso (PFd). Inicialmente pesquisado por Ochs et al. (1984), em grupo clínico restrito de pacientes com cistite intersticial, estudos ulteriores demonstraram sua alta freqüência na prática laboratorial em indivíduos com diversas condições não auto-imunes. De fato, este padrão é um dos mais freqüentes entre indivíduos não auto-imunes com reação FAN-HEp-2 positiva. A presente revisão tem como objetivo fornecer elementos que permitam ao clínico exercitar uma interpretação crítica dos resultados de FAN-HEp-2, incluindo a análise do título e do padrão de fluorescência.

Determination of autoantibodies against cellular antigens (ANA-HEp-2) has undergone several methodological changes over the decades and for an appropriate evaluation of results requires reflection on its sensitivity and specificity properties. Currently the test is known to have high sensitivity and questionable specificity since it is frequently positive in a considerable number of non-autoimmune subjects. Awareness that a substantial portion of the population has a positive ANA-HEp-2 test demands that characteristics permitting to distinguish positive tests in autoimmune patients from positive tests in non-autoimmune subjects be identified. Among these characteristics are noteworthy the titer and the fluorescence pattern. In particular, one must bear in mind that the fluorescence pattern results from recognition by autoantibodies of distinct cellular structures and, therefore, the fluorescence pattern provides a preliminary indication of possible autoantigens targeted by the sera being tested. An emblematic example is the nuclear dense fine speckled pattern. It was originally studied by Ochs and collaborators in 1994 in a restricted number of patients with interstitial cystitis. Further studies demonstrated its frequent occurrence in laboratory practice in individuals with diverse non-autoimmune clinical conditions. Indeed, this pattern is one of the most frequently observed in non-autoimmune patients with an ANA-HEp-2 positive reaction. The present review intends to provide elements allowing the clinician to perform a critical interpretation of results of an ANA-HE-2 test, including analysis of the titer and the fluorescence pattern.

65431 downloads Cited 1 time in SciELO

12.

Análise crítica do teste de anticorpos antinúcleo (FAn) na prática clínica
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Dellavance, Alessandra

; Leser, Paulo Guilherme

; Andrade, Luís Eduardo Coelho

Revista Brasileira de Reumatologia

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aug 2007, Volume 47 N. 4 Pages 265 - 275

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

A pesquisa de anticorpos contra antígenos celulares, tradicionalmente denominada FAN (fator antinúcleo) ou FAN-HEp-2, tem apresentado contínua evolução que requer dos profissionais envolvidos uma acurada e constante revisão sobre os paradigmas que norteiam a interpretação dos resultados obtidos. Os avanços metodológicos ocasionaram expressivo aumento na sensibilidade do teste e conseqüente diminuição de sua especificidade. Isto se verifica pelo crescente número de exames positivos em indivíduos aparentemente hígidos. Este fato tem criado situações desconfortáveis e muitas vezes angustiantes, sendo necessário buscar elementos para auxiliar os profissionais envolvidos. Estudos criteriosos e a experiência têm nos ensinado que há características peculiares usualmente associadas aos auto-anticorpos observados em pacientes auto-imunes e em indivíduos não auto-imunes. A presente revisão fornece uma abordagem sobre os pontos importantes para a correta valorização dos achados do teste do FAN-HEp-2 e que possam auxiliar na identificação de pacientes com doenças auto-imunes. Discute-se a importância do título e do padrão de imunofluorescência na valorização de um teste positivo e no desdobramento da pesquisa de auto-anticorpos específicos. Indivíduos não auto-imunes usualmente apresentam FAN-HEp-2 em títulos baixos, enquanto pacientes auto-imunes, geralmente, títulos médios ou altos. Entretanto, observam-se freqüentes exceções em ambos os contextos. Os padrões de fluorescência nucleares pontilhado grosso e homogêneo quase sempre se associam a um contexto de auto-imunidade sistêmica. Por outro lado, o padrão nuclear pontilhado fino denso é um dos mais freqüentemente observados em pessoas não auto-imunes com exame positivo de FAN-HEp-2. O Consenso Nacional para Padronização dos Laudos de FAN-HEp-2 tem desempenhado importante papel na melhoria da interpretação do exame de FAN-HEp-2 em nosso país. Finalmente, há que se considerar os possíveis e diversos significados de um teste positivo de FAN-HEp-2 em um paciente sem evidência objetiva de doença auto-imune.

Due to its continuous evolving process the fluorescent antinuclear antibody test (ANA-HEp-2) has required constant update efforts from personal involved in performing and interpreting its results. The methodological advances have brought up a considerable improvement in the test's sensitivity and consequently a decrease in its specificity. This has resulted in an increasing number of positive tests in apparently healthy subjects. This fact contributes to embarrassing situations and implies in the necessity to seek for elements to help the involved professionals. Well-conducted studies and experience have shown that there are some peculiar features associated with the auto-antibodies in patients with autoimmune diseases that are not present in those observed in healthy subjects. The present review brings an approach on the most important points to be considered in the analysis and evaluation of an ANA test that might help the identification of patients with autoimmune disease. Titer and immunofluorescence pattern are important parameters in the evaluation of the significance of a positive ANA-HEp2 test and in the reflex demand for further tests for specific autoantibodies. In general non-autoimmune individuals present low titer ANA-HEp-2 and patients with systemic autoimmune diseases present moder ate or high titer ANA-HEp-2 tests. However, exceptions to this are normally observed in both contexts. The homogeneous and coarse speckled nuclear patterns are almost exclusively observed in patients with systemic autoimmunity. On the other hand, the nuclear dense fine speckled pattern is one of the most frequently observed patterns in non-autoimmune individuals with a positive ANA-HEp-2 test. The basic concepts of the National Consensus on Standardization of ANA-HEp-2 Report have helped Brazilian rheumatologists and pathologists in the correct interpretation of the great diversity of ANA-HEp-2 immunofluorescence patterns. Finally, it is important to consider the possible and alternative significance of a positive ANA test in patients without objective evidence of autoimmune diseases.

92390 downloads Cited 7 times in SciELO

13.

Como interpretar e valorizar adequadamente o teste de anticorpos antinúcleo
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Dellavance, Alessandra

; Andrade, Luís Eduardo Coelho

Jornal Brasileiro de Patologia e Medicina Laboratorial

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jun 2007, Volume 43 N. 3 Pages 157 - 168

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

Atualmente, a pesquisa de anticorpos contra antígenos celulares, historicamente chamados de fator antinúcleo (FAN), tem requerido dos profissionais que trabalham (direta ou indiretamente) com o exame uma acurada e constante revisão dos paradigmas que norteiam a interpretação dos resultados obtidos. Os avanços metodológicos levaram a um aumento expressivo na sensibilidade do teste e, conseqüentemente, à diminuição de sua especificidade, fato comprovado pelo crescente número de exames positivos em indivíduos sabidamente hígidos. Há características peculiares, as quais usualmente são associadas aos auto-anticorpos observados em pacientes auto-imunes e em indivíduos não-auto-imunes. O objetivo desta revisão é trazer uma abordagem sobre os pontos importantes, para a correta valorização dos achados do teste do FAN, que possa auxiliar na identificação de pacientes com doenças auto-imunes. É discutida a importância do título e do padrão de imunofluorescência na valorização de um teste positivo e no desdobramento da pesquisa de auto-anticorpos específicos. Também é feita uma revisão dos conceitos do Consenso Nacional para Padronização dos Laudos de FAN-HEp-2. Finalmente, é oferecida uma discussão sobre os possíveis significados do teste positivo de FAN em um paciente sem evidência objetiva de doença auto-imune.

The tradicional fluorescent antinuclear antibody test (ANA) has required constant update efforts from personel involved in performing and interpreting its results. The methodological advances have brought up a considerable improvement in the test’s sensitivity and, consequentely, a decrease in its specificity. This has resulted in an increasing number of positive tests in apparently healthy subjects. However, there are some peculiar features associated with the auto-antibodies in patients with autoimmune diseases that are not present in those observed in healthy subjects. The objective of the present review is to bring an approach on the most important points to be considered in the analysis and evalution of an ANA test that might help in the identification of patients with autoimmune disease. Title and immunofluorescence pattern are discussed as important parameters and they are important in the evaluation of ANA and in the reflex demand for further tests for specific autoantibodies. The basic concepts of the National Consensus on Standardization of ANA-HEp-2 Report are posted. Finally, we explore the possible meanings of a positive ANA test in a patient without objective evidence of autoimmune disease.

76020 downloads Cited 5 times in SciELO

14.

II Consenso Brasileiro de Fator Antinuclear em Células HEp-2: Definitions for standardization of autoantibody testing against the nucleus (ANA HEp-2), nucleolus, cytoplasm and mitotic apparatus, as wel as its clinical associations
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Dellavance, Alessandra

; Gabriel Júnior, Alexandre

; Cintra, Alice F. U.

; Ximenes, Antônio Carlos

; Nuccitelli, Barbara

; Taliberti, Ben Hur

; Moreira, Caio

; von Mühlen, Carlos Alberto

; Bichara, Carlos David

; Santos, Cláudio Henrique Ramos dos

; Yano, Cristiane Martinez

; Mangueira, Cristóvão Luis Pitangueira

; Carvalho, Darlene Gonçalves

; Bonfá, Eloísa Silva Dutra de O.

; Doi, Elvira M.

; Guimarães, Fabiana Nunes de Carvalho

; Araújo, Flávia Ikeda e

; Mundim, Hugo Mendonça

; Rego, Jozelia

; Vieira, Lisiane Ericonio dos Anjos

; Poli, Luciana

; Andrade, Luís Eduardo Coelho

; Callado, Maria Roseli

; Mesquita, Mauro Meira

; Sugiyama, Mitiko

; Slhessarenko, Natasha

; Silva, Nilzio Antônio da

; Carballo, Orlando Gabriel

; Leser, Paulo Guilherme

; Francescantonio, Paulo Luiz Carvalho

; Jarach, Renata

; Xavier, Ricardo Machado

; Levy, Roger Abramino

; Neves, Suzane P. F.

; Cruvinel, Wilson de Melo

; Santos, Wilton Silva dos

Revista Brasileira de Reumatologia

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jun 2003, Volume 43 N. 3 Pages 129 - 140

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

OBJETIVO: O Segundo Consenso Brasileiro de Fator Antinuclear (FAN) em Células HEp-2 ratificou os algoritmos de decisão para leitura dos padrões do FAN na imunofluorescência indireta vistos na primeira edição do Consenso Brasileiro, adicionando ainda um novo algoritmo relacionado com os padrões mistos. MÉTODOS: Tendo em vista a habilidade do teste em detectar autoantígenos nos distintos compartimentos celulares, e não apenas no núcleo, propõe-se novas denominações para este exame laboratorial. RESULTADOS E CONCLUSÕES: Como novas denominações algumas sugestões foram igualmente aceitas, dentro do tema "pesquisa de auto-anticorpos contra constituintes do núcleo (FAN HEp-2), nucléolo, citoplasma e aparelho mitótico". Foram abordadas as principais relevâncias clínicas com os padrões de FAN descritos, facilitando o melhor uso do ensaio pelo médico.

OBJECTIVE: The Second Brazilian Consensus on Antinuclear Antibodies (ANA) in HEp-2 Cells approved and extended the decision trees developed during the First Brazilian Consensus in order to also offer information about mixed patterns of fluorescence. METHODS: Since this test elicits reactions not only to nuclear autoimmune antigens but also to different cell compartments, new denominations for the test were approved. Results and CONCLUSIONS: These new denominations encompass variations on the "autoantibody testing against the nucleus (ANA HEp-2), nucleolus, cytoplasm, and mitotic apparatus" issue. Furthermore, major clinical associations were described for each immunofluorescent pattern, facilitating the interpretation of laboratory results in the clinical practice.

18662 downloads

15.

I Consenso Nacional para Padronização dos Laudos de FAN HEp-2
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Dellavance, Alessandra

; G. Júnior, Alexandre

; Cintra, Alice F.U.

; Ximenes, Antônio C.

; Nuccitelli, Barbara

; Mühlen, Carlos A. von

; Bichara, Carlos D.

; Yano, Cristiane

; Carvalho, Darlene G.

; Bonfá, Eloísa S.D.O.

; Guimarães, Fabiana N.C.

; Mundim, Hugo M.

; Pfrimer, Irmtraut A.H.

; Rego, Jozelia

; Andrade, Luiz E.C.

; Mesquita, Mauro M.

; Santiago, Mittermayer B.

; Silva, Nilzio A.

; Miranda, Paulo J.

; Leser, Paulo

; Francescantonio, Paulo Luiz C.

; Jarach, Renata

; Levy, Roger A.

; Neves, Suzane P.F.

; Cruvinel, Wilson M.

; Santos, Wilton S.

Jornal Brasileiro de Patologia e Medicina Laboratorial

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jul 2002, Volume 38 N. 3 Pages 207 - 216

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

A análise da presença de auto-anticorpos feita por imunofluorescência indireta em células HEp-2 constitui-se em um método de triagem escolhido na maioria dos laboratórios clínicos. A ausência de uma nomenclatura definida para a descrição dos laudos tem trazido problemas na utilização clínica do teste, pelas dificuldades no controle de qualidade e na padronização dos resultados, que, por sua vez, embora similares, recebiam denominações diferentes. O I Consenso Brasileiro para Padronização dos Laudos de FAN HEp-2 reuniu em agosto de 2000, em Goiânia, diversos especialistas de todo o Brasil. Esses emitiram pareceres em consenso para os distintos padrões: nucleares, nucleolares, citoplasmáticos e aparelho mitótico. Foram feitas recomendações sobre os critérios para a leitura de uma lâmina, bem como para relação entre a diluição de triagem e o sistema óptico utilizado.

The technique of immunofluorescence using HEp-2 cells as substrate is the screening method of choice for the presence of autoantibodies in many clinical laboratories. The lack of a specific terminology for reporting results brings problems in quality control, clinical utility of the test, and standardization attempts. The first Brazilian Consensus for Standardization of ANA in HEp-2 Cells took place in Goiânia in August 2000. Several laboratory specialists with experience in the methodology showed up. They established guidelines for the description of ANA patterns in the Portuguese language, encompassing distinct descriptions for nuclear, nucleolar, cytoplasmic and mitotic apparatus patterns of fluorescence. Recommendations were also established regarding screening titers, final dilution titer, and on morphological criteria for reading the slides.

11489 downloads Cited 5 times in SciELO

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