title: Normal lymphocytes from leukemic samples as an internal quality control for fluorescence intensity in immunophenotyping of acute leukemias.
abstract: BACKGROUND
  Multiparametric flow cytometry has become an indispensable but complex tool for the diagnosis of acute leukemias. Interpretation of immunophenotypic data within a six-parameter analytical space relies on the standardization and validation of the instrument, the reagents, and the procedure. To address whether or not residual normal lymphocytes, usually present within leukemic samples, can serve as internal quality control for fluorescence intensity, 116 leukemic and 35 normal samples were analyzed.
  
  
  METHODS
  Eight laboratories participated in the study and recruited a total of 151 individuals including 29 patients with B-cell precursor acute lymphoblastic leukemia (BCP-ALL), 77 with acute myeloid leukemia (AML), 10 with T-cell precursor acute lymphoblastic leukemia (T-ALL), and 35 normal bone marrow donors. Lymphocytes were gated according to the CD45hi/SSClo gating strategy, after which median fluorescence intensities (MFI) as well as percentages of positive cells (%positive) for CD19, CD22, CD7, and CD3 were recorded. Nonparametric statistics were used to compare variation within and between laboratories.
  
  
  RESULTS
  Normal lymphocytes within leukemic samples do not show substantial differences compared to lymphocytes from normal controls with respect to expression of CD19, CD22, CD7, and CD3. In particular, longitudinal control charts of MFI values for CD3 antigen provide useful information on analytical and instrument performance.
  
  
  CONCLUSIONS
  Residual normal lymphocytes can serve as internal quality control for studies addressing fluorescence intensity in the setting of immunophenotyping of acute leukemias.
authors.affiliation: HELIOS Klinikum Berlin, Charité-University Medicine Berlin, Germany. rratei@berlin.helios-kliniken.de
authors.forename: Richard
authors.initials: R
authors.lastname: Ratei
authors.forename: Leonid
authors.initials: L
authors.lastname: Karawajew
authors.forename: Francis
authors.initials: F
authors.lastname: Lacombe
authors.forename: Krystyna
authors.initials: K
authors.lastname: Jagoda
authors.forename: Giovanni
authors.initials: G
authors.lastname: Del Poeta
authors.forename: Jaco
authors.initials: J
authors.lastname: Kraan
authors.forename: Maria
authors.initials: M
authors.lastname: De Santiago
authors.forename: János
authors.initials: J
authors.lastname: Kappelmayer
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Björklund
authors.forename: W-D
authors.initials: WD
authors.lastname: Ludwig
authors.forename: Jan
authors.initials: J
authors.lastname: Gratama
authors.forename: Alberto
authors.initials: A
authors.lastname: Orfao
mesh_terms: D000208:Acute Disease; D016022:Case-Control Studies; D005434:Flow Cytometry / Q000379:methods*; D005453:Fluorescence; D006801:Humans; D016130:Immunophenotyping / Q000379:methods*; D007938:Leukemia / Q000175:diagnosis; D008214:Lymphocytes / Q000166:cytology* / Q000378:metabolism*; D011786:Quality Control; D012015:Reference Standards