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Published on: October 19, 2014
Normal lymphocytes from leukemic samples as an internal quality control for fluorescence intensity in
Richard Ratei1, Leonid Karawajew, Francis Lacombe
1HELIOS Klinikum Berlin, Charité-University Medicine Berlin, Germany. rratei@berlin.helios-kliniken.de
Insights
Residual normal lymphocytes in acute leukemia samples can act as internal quality control for fluorescence intensity measurements, ensuring reliable immunophenotyping. This method aids in standardizing diagnostic procedures for better accuracy.
Area of Science:
- Hematology
- Immunology
- Clinical Diagnostics
Background:
- Multiparametric flow cytometry is crucial for acute leukemia diagnosis but requires standardization.
- Interpreting immunophenotypic data necessitates validated instruments, reagents, and procedures.
- The role of residual normal lymphocytes as internal quality control for fluorescence intensity is under investigation.
Purpose of the Study:
- To evaluate if residual normal lymphocytes in acute leukemia samples can serve as internal quality control for fluorescence intensity.
- To assess the reliability of immunophenotypic data through internal standardization.
Main Methods:
- Analyzed 116 leukemic and 35 normal bone marrow samples across eight laboratories.
- Gated lymphocytes using CD45hi/SSClo strategy and recorded median fluorescence intensities (MFI) and %positive for CD19, CD22, CD7, and CD3.
- Employed nonparametric statistics to compare variations within and between laboratories.
Main Results:
- Normal lymphocytes in leukemic samples exhibited similar expression of CD19, CD22, CD7, and CD3 compared to normal controls.
- Longitudinal control charts of MFI for CD3 antigen demonstrated utility in monitoring analytical and instrument performance.
- No substantial differences were observed in antigen expression between residual normal lymphocytes and control lymphocytes.
Conclusions:
- Residual normal lymphocytes are suitable for internal quality control in fluorescence intensity studies for acute leukemia immunophenotyping.
- This approach enhances the standardization and validation of flow cytometry procedures in leukemia diagnostics.
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.
Conclusion:
Residual normal lymphocytes can serve as internal quality control for studies addressing fluorescence intensity in the setting of immunophenotyping of acute leukemias.
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