Standardization of Flow Cytometric Immunophenotyping for Hematological Malignancies: The FranceFlow Group Experience.

Flow cytometry is broadly used for the identification, characterization, and monitoring of hematological malignancies. However, the use of clinical flow cytometry is restricted by its lack of reproducibility across multiple centers. Since 2006, the EuroFlow consortium has been developing a standardized procedure detailing the whole process from instrument settings to data analysis. The FranceFlow group was created in 2010 with the intention to educate participating centers in France about the standardized instrument setting protocol (SOP) developed by the EuroFlow consortium and to organise several rounds of quality controls (QCs) in order to evaluate the feasibility of its application and its results. Here, we report the 5 year experience of the FranceFlow group and the results of the seven QCs of 23 instruments, involving up to 19 centers, in France and in Belgium. The FranceFlow group demonstrates that both the distribution and applicability of the SOP have been successful. Intercenter reproducibility was evaluated using both normal and pathological blood samples. Coefficients of variation (CVs) across the centers were <7% for the percentages of cell subsets and <30% for the median fluorescence intensities (MFIs) of the markers tested. Intracenter reproducibility provided similar results with CVs of <3% for the percentages of the majority of cell subsets, and CVs of <20% for the MFI values for the majority of markers. Altogether, the FranceFlow group show that the 19 participating labs might be considered as one unique laboratory with 23 identical flow cytometers able to reproduce identical results. Therefore, SOP significantly improves reproducibility of clinical flow in hematology and opens new avenues by providing a robust companion diagnostic tool for clinical trials in hematology. © 2019 International Society for Advancement of Cytometry.

Cytomorphology and flow cytometry of brain biopsy rinse fluid enables faster and multidisciplinary diagnosis of large B-cell lymphoma of the central nervous system.

BACKGROUND: Central nervous system lymphomas are aggressive tumors requiring a prompt diagnosis for successful treatment. Stereotactic biopsy remains the standard procedure, but the time needed for histopathology is usually over 2 days. We evaluated the contribution of cytomorphology and flow cytometry to histopathology of the brain biopsy in particular on the rinse fluid usually removed. METHODS: Eighteen patients with suspected localized brain lymphoma underwent stereotactic brain biopsy. Brain biopsy tissue sample and/or brain biopsy rinse fluid were analyzed by cytomorphology combined with flow cytometry. Histopathology was used as a reference. RESULTS: Histopathology characterized ten diffuse large B-cell lymphomas and eight other diseases. Cytomorphology and flow cytometry showed lymphoma cells in nine out of the ten lymphomas. Three cytomorphology or flow cytometry negative results were reported for lymphomas in tissue samples due to low cellularity and biopsy sample conditioning. No lymphomatous cells were found by cytomorphology or flow cytometry in the eight other diseases. Rinse fluid results were consistent with histology in all cases studied (sensitivity and specificity, 100%). The median time to result was 4.5 days (range, 2-10 days) for histopathology, while 5 h (range, 3-20 h) were required for both cytomorphology and flow cytometry. CONCLUSIONS: Brain biopsy rinse fluid alleviates problems of tissue sample distribution compared to tissue sample. Its analysis performs the diagnosis of B-cell lymphoma in a few hours and, associated with histopathology, allows a multidisciplinary diagnosis. This study shows that cytomorphology combined with flow cytometry on brain biopsy rinse fluid is a new, fast, and useful strategy. © 2016 International Clinical Cytometry Society.

A case report of pseudo grey platelet syndrome with citrate-induced pseudothrombocytopenia: those artifacts may interfere in the platelet numeration and lead to critical misdiagnosis.

The pseudo grey platelet syndrome is a rare artifact due to the degranulation of platelets caused, in vitro, by EDTA. This phenomenon is likely to disturb the platelet numeration and it is essential not to mistake it for a grey syndrome platelet, which is a constitutional thrombopathy with macrothrombopenia, in order to avoid specialized tests, or even misdiagnosis. Indeed, these two entities are cytologically alike, as grey platelets are found on the blood smear of a sample collected on EDTA in both cases. We here describe the case of a patient admitted in Colmar's Hospital for a chronic thrombocytopenia, associating both a pseudothrombocytopenia and a pseudo grey platelet syndrome.

[Comparison of routine use of two chromogenic media ChromID CPS (bioMérieux) and UriSelect4 (Bio-Rad) for the detection of Escherichia coli and major uropathogenics in urine]. / Comparaison de l'utilisation en routine de deux milieux chromogènes ChromID CPS (bioMérieux) et UriSelect4 (Bio-Rad) pour la détection d'Escherichia coli et des principaux uropathogènes dans les urines.

Escherichia coli is the most common bacterial cause of urinary tract infections. Its rapid and specific identification in urine samples represents a major challenge within the rendering results and optimizing the management of the patient. We aimed to compare the sensitivity and specificity of two commercially available chromogenic media for E. coli: ChromID CPS (Biomérieux) and UriSelect4 (Bio(-)Rad), without carrying out further tests. 99 consecutive and non-redundant urine samples considered to be infected were simultaneously plated onto blood agar and the two chromogenic media. Colony color and bacterial growth quantification were compared 18 and 48 hours after incubation. Bacteria were identified with mass spectrometry. A complementary analysis on 80 bacterial strains known to pose potential identification problems was performed. 43 urines samples grew E. coli, and 42 of them were pink-colored on the two chromogenic mediums, as expected (sensibility=97.7%). Growth quantification was significantly greater on blood agar than on chromogenic media (p<0.001).We noted specificity issues at the complementary analysis with the UriSelect4 medium: Citrobacter freundii and some strains of Citrobacter brakii, Enterobacter cloacae and Hafnia alvei were pink-colored, and could be misidentified as E. coli. ChromID CPS medium did not show such misidentification. In conclusion, the agar ChromID CPS proved to be greater than the UriSelect4 agar in our work in terms of specificity of direct identification of E. Coli, without the use of additional test.