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Cytometry B Clin Cytom. 2018 Jan;94(1):82-93. doi: 10.1002/cyto.b.21518. Epub 2017 Feb 21.

AIEOP-BFM consensus guidelines 2016 for flow cytometric immunophenotyping of Pediatric acute lymphoblastic leukemia.

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Children's Cancer Research Institute and St. Anna Children's Hospital, Department of Pediatrics, Medical University of Vienna, Vienna, Austria.
Laboratory of Pediatric Onco-Hematology, Women and Child Department, University of Padova, Padova, Italy.
Tettamanti Research Center and Department of Pediatrics, Ospedale San Gerardo, University of Milano-Bicocca, Monza, Italy.
Clinic for Oncology and Tumor Immunology, HELIOS Klinikum Berlin-Buch, Berlin, Germany.
Department of Pediatric Hematology and Oncology, 2nd Faculty of Medicine, Charles University and University Hospital Motol, Prague, Czech Republic.
Department of Pediatric Hematology and Oncology, Schneider's Children's Medical Center, Petach-Tikva, Israel.
Cancer Research Center and the Hematology Laboratory, Jeffrey Modell Foundation (JMF) Center, Edmond and Lily Safra Children's Hospital, Sheba Medical Center, Tel Hashomer, Israel.
Division of Oncology and Children's Research Center, University Children's Hospital, University of Zurich, Zurich, Switzerland.
Flow Cytometry Unit, Institute of Clinical Pathology and Medical Research, Westmead Hospital, Sydney, Australia.
Department of Pediatric Oncology/Hematology, Charité Universitätsmedizin, Berlin, Germany.
Department of Pediatrics, University Medical Center Schleswig-Holstein, Kiel, Germany.


Immunophenotyping by flow cytometry (FCM) is a worldwide mainstay in leukemia diagnostics. For concordant multicentric application, however, a gap exists between available classification systems, technologic standardization, and clinical needs. The AIEOP-BFM consortium induced an extensive standardization and validation effort between its nine national reference laboratories collaborating in immunophenotyping of pediatric acute lymphoblastic leukemia (ALL). We elaborated common guidelines which take advantage of the possibilities of multi-color FCM: marker panel requirements, immunological blast gating, in-sample controls, tri-partite antigen expression rating (negative vs. weak or strong positive) with capturing of blast cell heterogeneities and subclone formation, refined ALL subclassification, and a dominant lineage assignment algorithm able to distinguish "simple" from bilineal/"complex" mixed phenotype acute leukemia (MPAL) cases, which is essential for choice of treatment. These guidelines are a first step toward necessary inter-laboratory standardization of pediatric leukemia immunophenotyping for a concordant multicentric application.


flow cytometry; immunophenotyping:; leukemia; pediatric; standardization

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