ABSTRACT Introduction: Flow cytometry has become an increasingly important tool in the clinical laboratory for the diagnosis and monitoring of many hematopoietic neoplasms. This method is ideal for immunophenotypic identification of cellular subpopulations in complex samples, such as bone marrow and peripheral blood. In general, 4-color panels appear to be adequate, depending on the assay. In acute leukemias (ALs), it is necessary identify and characterize the population of abnormal cells in order to recognize the compromised lineage and classify leukemia according to the WHO criteria. Although the use of eightto ten-color immunophenotyping panels is wellestablished, many laboratories do not have access to this technology. Objective and Method: In 2015, the Brazilian Group of Flow Cytometry (Grupo Brasileiro de Citometria de Fluxo, GBCFLUX) proposed antibody panels designed to allow the precise diagnosis and characterization of AL within available resources. As many Brazilian flow cytometry laboratories use four-color immunophenotyping, the GBCFLUX has updated that document, according to current leukemia knowledge and after a forum of discussion and validation of antibody panels. Results: Recommendations for morphological analysis of bone marrow smears and performing screening panel for lineage (s) identification of AL were maintained from the previous publication. The lineage-oriented proposed panels for B and T cell acute lymphoblastic leukemia (ALL) and for acute myeloid leukemia (AML) were constructed for an appropriate leukemia classification. Conclusion: Three levels of recommendations (i.e., mandatory, recommended, and optional) were established to enable an accurate diagnosis with some flexibility, considering local laboratory resources and patient-specific needs.

ABSTRACT Leishmaniasis is a disease complex with various clinical symptoms caused by different species of parasites of the genus Leishmania. The visceral form of the disease, characterized by severe symptoms is fatal, if not treated. The high toxicity of current antileishmanial drugs and the need for long-term treatment make the therapy complicated, especially in a large number of infected children. Hence, the search for new therapies must be intensified. Oral administration of the trace element zinc has been considered in alternative treatments against different clinical forms of leishmaniasis. This study revealed that the administration of zinc in children with visceral leishmaniasis, during treatment with amphotericin B or glucantime, accelerates the regression of the spleen enlargement without interfering with the recovery of hematological parameters.

ABSTRACT When the FLT3 gene is mutated, it originates a modified receptor with structural changes, which give survival advantage and malignant hematopoietic cell proliferation. Thus, the presence of mutations in this gene is considered an unfavorable prognostic factor. A total of 85 consecutive samples of newly diagnosed untreated patients with AL were included in the study after they provided their informed consent. FLT3 gene mutations were detected by PCR. For the pediatric group, a positive correlation was observed between WBC count and the presence of FLT3-ITD in patients with AML and ALL. Furthermore, children with AML who had the FLT3-ITD mutation showed a tendency to express CD34 in blast cells. In the adult group, the AML patients with FLT3-ITD who expressed CD34 in blast cells had a tendency to worse progression. The present data indicate no association between the prognostic factors evaluated and FLT3 gene mutations in adult with AL. Yet, the presence of FLT3-ITD mutation was significantly related with WBC count in the pediatric group. These findings demonstrate that FLT3 gene mutations can be considered as independent poor prognostic factors.

Chronic myeloproliferative neoplasms arise from clonal proliferation of hematopoietic stem cells. According to the World Health Organization myeloproliferative neoplasms are classified as: chronic myelogenous leukemia, polycythemia vera, essential thrombocythemia, primary myelofibrosis, chronic neutrophilic leukemia, chronic eosinophilic leukemia, hypereosinophilic syndrome, mast cell disease, and unclassifiable myeloproliferative neoplasms. In the revised 2008 WHO diagnostic criteria for myeloproliferative neoplasms, mutation screening for JAK2V617F is considered a major criterion for polycythemia vera diagnosis and also for essential thrombocythemia and primary myelofibrosis, the presence of this mutation represents a clonal marker. There are currently two hypotheses explaining the role of the JAK2V617F mutation in chronic myeloproliferative neoplasms. According to these theories, the mutation plays either a primary or secondary role in disease development. The discovery of the JAK2V617F mutation has been essential in understanding the genetic basis of chronic myeloproliferative neoplasms, providing some idea on how a single mutation can result in three different chronic myeloproliferative neoplasm phenotypes. But there are still some issues to be clarified. Thus, studies are still needed to determine specific molecular markers for each subtype of chronic myeloproliferative neoplasm.

As leucemias mieloides agudas (LMA) constituem um grupo de neoplasias malignas caracterizadas pela proliferação descontrolada de células hematopoéticas, decorrente de mutações que podem ocorrer em diferentes fases da diferenciação de células precursoras mieloides. Em 2008, a Organização Mundial da Saúde (OMS-2008) publicou uma nova classificação para neoplasias do sistema hematopoético e linfoide. De acordo com essa classificação, para um diagnóstico mais preciso e estratificação de prognóstico de pacientes com leucemias mieloides agudas, devem-se pesquisar mutações nos genes FLT3 e NPM1. Sabe-se que a presença de mutações no gene FLT3 é de prognóstico desfavorável e que as mutações no gene NPM1 do tipo A são de prognóstico favorável. Assim, nos países desenvolvidos, a análise das mutações no gene FLT3 e NPM1 tem sido considerada como um fator de prognóstico importante na decisão terapêutica em pacientes com diagnóstico de leucemias mieloides agudas. Considerando essas informações, é de extrema importância a análise das mutações no gene FLT3 (duplicação interna em tandem - DIT - e mutação pontual D835) e no gene NPM1 como marcadores moleculares para o diagnóstico, o prognóstico e a monitoração de doença residual mínima em pacientes com leucemias mieloides agudas.

Acute myeloid leukemia (AML) is a group of malignancies characterized by uncontrolled proliferation of hematopoietic cells resulting from mutations that occur at different stages in the differentiation of myeloid precursor cells. In 2008, the World Health Organization (WHO-2008) published a new classification for cancers of the hematopoietic and lymphoid system. According to this classification, FLT3 and NPM1 gene mutations should be investigated for a more precise diagnosis and prognostic stratification of AML patients. It is well known that the presence of FLT3 gene mutations is considered an unfavorable prognostic factor and type-A NPM1 gene mutations are considered to be favorable. In developed countries, an analysis of FLT3 and NPM1 mutations is considered important for therapeutic decisions in AML patients. Hence, an analysis of FLT3 (internal tandem duplication - ITD- and D835 point mutation) and NPM1 gene mutations is extremely important as molecular markers for diagnosis, prognosis and monitoring of minimal residual disease in LMA patients.

O linfoma de Hodgkin (LH) e o linfoma difuso de grandes células B (LDGCB) são neoplasias derivadas de células B, que apresentam marcadores imunofenotípicos em comum, e, por isso, em alguns casos onde ocorre sobreposição morfológica torna-se difícil o diagnóstico diferencial entre eles. O grau de dificuldade aumenta quando o tumor é localizado no mediastino. O fato de a biópsia de mediastino ser um procedimento ainda mais invasivo que a maioria das outras biópsias, muitas vezes faz com que sejam coletadas quantidades insuficientes de amostra, o que dificulta a análise das características morfológicas da linfoproliferação. Considerando tais dificuldades, neste trabalho investigaremos a expressão imuno-histoquímica da proteína Pax-5 no diagnóstico diferencial de LH e LDGCB. Foram analisadas amostras de 14 pacientes, sendo dez com LDGCB e quatro com LH. O estudo imuno-histoquímico com Pax-5 mostrou 100% de positividade nuclear para LDGCB. Porém, destes, 30% mostraram positividade forte (+++); 50% moderada (++); 20% fraca (+). Já nas amostras de LH, 50% obtiveram marcação focal e fraca nas células de Reed/Sternberg (CRS) e 50% foram negativas. A marcação focal e fraca, ou até negativa, do Pax-5 em LH pode ser um ponto importante para o diagnóstico diferencial, uma vez que difere muito da marcação visualizada no LDGCB. Assim, é possível concluir que o anticorpo Pax-5 é um bom marcador para ser usado em conjunto com outros marcadores no painel de rotina e, quando associado à clínica, pode ser muito útil no diagnóstico diferencial entre LDGCB e LH em linfoma de mediastino.

Hodgkin's Lymphoma (HL) and diffuse large B-cell Lymphoma (DLBCL) are neoplasias derived from B cells that present the same immunophenotypic markers, making the differential diagnosis between them very difficult particularly in cases where morphological overlapping occurs. An extra difficulty exists when the tumor is located in the mediastinum, thereby demanding a more invasive biopsy. In addition, frequently an excessively small sample is collected, putting the analysis of the morphological characteristics of lymphoproliferation at risk. The goal of this study was to evaluate PAX-5 as a marker to distinguish HL from DLBCL. Overall, 10 out of 14 studied cases had a diagnosis of DLBCL and 4 patients of HL. The Pax-5 immunohistochemical staining showed 100% positivity in the LBCL study group. However, 30% showed strong positivity (+++), 50% moderate (++) and 20% weak (+). On the other hand, 50% of the HL group showed a weak focal pattern of staining of the Reed/Sternberg Cells (RSC), whereas the other 50% showed negativity. The weak focal pattern as well as the negative staining pattern of PAX-5 in HL may be an important key in the differential diagnosis as it is very different from DLBCL. Hence, we suggest that the PAX-5 antibody is a good immunomarker and may be used with other markers in the laboratory routine. It is also very useful when associated to clinical features of patients to reach a differential diagnosis between DLBCL and HL mediastinal lymphoma.

As síndromes mielodisplásicas (SMDs) são caracterizadas por uma desordem clonal de células primordiais (stem cell) e hematopoese ineficaz que levam à displasia de uma ou mais linhagens celulares da medula óssea, citopenias periféricas e instabilidade genética, as quais aumentam o risco de transformação à leucemia mieloide aguda. Esse grupo heterogêneo de doenças hematopoéticas pode surgir como doença primária, que possui etiologia variada e não completamente definida, ou secundária ao tratamento quimioterápico ou radioterápico para outras neoplasias. O surgimento e aprimoramento de tecnologias de diagnóstico geraram uma melhor compreensão dos processos envolvidos na gênese e evolução das SMDs, o que possibilitou o desenvolvimento de marcadores de diagnóstico e acompanhamentos cada vez mais precoces e específicos. No ano de 2008, a Organização Mundial da Saúde (OMS) redefiniu os critérios para classificação das SMDs, dividindo-as em sete subgrupos. Nessa classificação foram incluídos novos aspectos imunofenotípicos, genéticos, citomorfológicos e moleculares, o que tornou o domínio e o acesso a tecnologias de ponta imprescindíveis para a realização do diagnóstico das SMDs. Apesar dos avanços tecnológicos, alguns pontos, como as bases moleculares da transformação de SMD para LMA, ainda não estão bem esclarecidos, fazendo necessária a continuação de estudos nessa área. Diante disso, essa revisão busca compilar dados atuais dos aspectos moleculares e laboratoriais das SMDs.

Myelodysplastic syndromes (MDSs) are characterized by a stem cell clonal disorder and ineffective hematopoiesis which causes dysplasia in one or more bone marrow hematopoietic cell lineages, peripheral cytopenia and genetic instability with enhanced risk to transform into acute myeloid leukemia (AML). This heterogeneous group of hematopoietic diseases can develop as primary diseases, which posses a variable and not completely defined etiology, or as secondary to chemotherapy or radiotherapy for other neoplasias. The evolution of diagnostic tests has improved comprehension of the process involved in the genesis and evolution of MDSs, making the development of earlier and more specific tests for diagnosis and follow ups possible. In 2008, the World Health Organization (WHO) redefined the criteria for the classification of MDSs, dividing them into seven subgroups. This classification included new immunophenotypic, genetic, cytomorphologic and molecular features, which are essential for the diagnosis of MDSs and for a better comprehension of the disease. Despite technological advances, some details, such as the molecular basis of the transformation of MDS to AML, are still not completely understood, which makes further studies in this field necessary. Hence, the objective of this review is to make a compilation of recent molecular and laboratory aspects of MDS.

O estudo avalia a capacidade do Therapeutic Intervention Scoring System (TISS-28) de discriminar pacientes internados em UTI, prováveis de morrer daqueles possíveis de sobreviver e estabelecer a pontuação limiar para alta probabilidade de morte. Os resultados, obtidos da amostra de 200 pacientes internados em 14 UTIs do Município de São Paulo, mostraram que o TISS-28 apresentou associação com mortalidade (p=0,0001). O ponto de corte estabelecido foi 21. Encontrou-se que 80,88% dos que morreram tinham pontuação do TISS-28 maior ou igual, e 68,18% dos sobreviventes tinham pontuação menor que 21. Além disso, quanto ao valor prognóstico do TISS-28, constatou-se acurácia de 0,72.

The study evaluates the competence of the "TISS-28" to distinguish inpatients at the ICU, between the ones likely to die from the ones likely to survive and to establish a threshold score for high likelihood to death. The findings obtained by the sample of 200 inpatients at 14 ICUs in Sao Paulo County showed that the TISS-28 presented association with mortality (p=0.0001). The cutting score established was 21. It was found that 80.88% of those who died had the TISS-28 score similar or higher and 68.18% of survivors had the score below 21. Furthermore, regarding to the prognostic value of TISS-28, it was showed up the accuracy of 0.72.

El estudio evalúa la capacidad del Therapeutic Intervention Scoring System (TISS-28) para discriminar pacientes internados en UCI, probables de morir de aquellos posibles de sobrevivir y establecer la puntuación límite para la alta probabilidad de muerte. Los resultados, obtenidos de la muestra de 200 pacientes internados en 14 UCIs del Municipio de São Paulo, mostraron que el TISS-28 presentó asociación con la mortalidad (p=0,0001). El punto de corte establecido fue 21. Se encontró que el 80.88% de los que murieron tenían puntuación del TISS-28 mayor o igual, y 68.18% de los sobrevivientes tenían puntuación menor que 21. Además de eso, en cuanto al valor pronóstico del TISS-28, se constató exactitud de 0.72.

The study evaluates the competence of the "TISS-28" to distinguish inpatients at the ICU, between the ones likely to die from the ones likely to survive and to establish a threshold score for high likelihood to death. The findings obtained by the sample of 200 inpatients at 14 ICUs in Sao Paulo County showed that the TISS-28 presented association with mortality (p=0.0001). The cutting score established was 21. It was found that 80.88% of those who died had the TISS-28 score similar or higher and 68.18% of survivors had the score below 21. Furthermore, regarding to the prognostic value of TISS-28, it was showed up the accuracy of 0.72.

El estudio evalúa la capacidad del Therapeutic Intervention Scoring System (TISS-28) para discriminar pacientes internados en UCI, probables de morir de aquellos posibles de sobrevivir y establecer la puntuación límite para la alta probabilidad de muerte. Los resultados, obtenidos de la muestra de 200 pacientes internados en 14 UCIs del Municipio de São Paulo, mostraron que el TISS-28 presentó asociación con la mortalidad (p=0,0001). El punto de corte establecido fue 21. Se encontró que el 80.88% de los que murieron tenían puntuación del TISS-28 mayor o igual, y 68.18% de los sobrevivientes tenían puntuación menor que 21. Además de eso, en cuanto al valor pronóstico del TISS-28, se constató exactitud de 0.72.

O estudo avalia a capacidade do Therapeutic Intervention Scoring System (TISS-28) de discriminar pacientes internados em UTI, prováveis de morrer daqueles possíveis de sobreviver e estabelecer a pontuação limiar para alta probabilidade de morte. Os resultados, obtidos da amostra de 200 pacientes internados em 14 UTIs do Município de São Paulo, mostraram que o TISS-28 apresentou associação com mortalidade (p=0,0001). O ponto de corte estabelecido foi 21. Encontrou-se que 80,88% dos que morreram tinham pontuação do TISS-28 maior ou igual, e 68,18% dos sobreviventes tinham pontuação menor que 21. Além disso, quanto ao valor prognóstico do TISS-28, constatou-se acurácia de 0,72.

Este estudo foi realizado em um ambulatório de transplante de medula óssea. A amostra constituiu-se de 17 pacientes e 6 funcionárias. A coleta de dados ocorreu no período de outubro à dezembro 1996, através de formulário, com decodificação de imagens filmadas, consulta de prontuários e de funcionárias. Os objetivos do estudo foram: identificar os tipos de toque e em que situações e regiões corpóreas ocorreram entre funcionárias e pacientes. Houve o predomínio do toque instrumental durante o atendimento, sendo mais freqüente durante o curativo de saída do cateter. As regiões mais tocadas foram a torácica e membros superiores.

This study was realized in the outpatient ward of bone marrow transplantation. The sample consisted of 17 adult patients and six workers. The data was collected from October to December, 1996, being obtained through filming, decodification of the films, a questionnaire and interview with the workers. The purposes were to identify in what situations the different types occurred, and which regions of the body were involved. It was found that most of the touching was of the instrumental type. The regions touched were the thorax and upper limbs.

This study was realized in the outpatient ward of bone marrow transplantation. The sample consisted of 17 adult patients and six workers. The data was collected from October to December, 1996, being obtained through filming, decodification of the films, a questionnaire and interview with the workers. The purposes were to identify in what situations the different types occurred, and which regions of the body were involved. It was found that most of the touching was of the instrumental type. The regions touched were the thorax and upper limbs.

Este estudo foi realizado em um ambulatório de transplante de medula óssea. A amostra constituiu-se de 17 pacientes e 6 funcionárias. A coleta de dados ocorreu no período de outubro à dezembro 1996, através de formulário, com decodificação de imagens filmadas, consulta de prontuários e de funcionárias. Os objetivos do estudo foram: identificar os tipos de toque e em que situações e regiões corpóreas ocorreram entre funcionárias e pacientes. Houve o predomínio do toque instrumental durante o atendimento, sendo mais freqüente durante o curativo de saída do cateter. As regiões mais tocadas foram a torácica e membros superiores.