| 1. |
- Björnsson, Sven, et al.
(författare)
-
Total nucleated cell differential for blood and bone marrow using a single tube in a five-color flow cytometer.
- 2008
-
Ingår i: Cytometry Part B - Clinical Cytometry. - : Wiley. - 1552-4949 .- 1552-4957. ; 74B, s. 91-103
-
Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND:: Flow cytometry allows the use of several antibodies in addition to light scatter, and most flow cytometers will provide at least seven measurements on each cell passing through the laser beam. A skilled microscopist will classify at least 14 cell classes in bone marrow or blood. Our goal was to use the seven parameters available in our flow cytometer to provide a reliable differential count using only one tube. METHODS:: Peripheral blood samples were analyzed on the Beckman Coulter LH750 cell counter, and the flagging and messages from the cell counter were used to select normal or pathological samples. Samples without flags (N = 50), with >2% erythroblasts (N = 80), or with "Blast" or "Verify diff" flags (N = 54) were investigated. We used a lyse-no-wash method to ensure minimal loss of fragile cells with live gating on DRAQ5-positive cells to acquire only nucleated cells. The FL-1 to FL-4 channels were used for the antibodies CD36-FITC, CD203-PE, CD138-PE, CD45-ECD, CD16-Pcy5, and CD56-Pcy5. FL-5 was used for the DNA-stain DRAQ5. RESULTS:: Using live gate acquisition on DRAQ5, we were able to classify total nucleated cells into 10 classes. We were unable to identify megakaryocytes, but platelets could be studied by rerunning the sample after dilution and gating on DRAQ5-negative CD36-posive events. Validation against digitized microscopy and cell counter showed linear correlations within each cell class with correlation coefficients that seem reasonable for cellular classification. The lowest correlation was found for basophil granulocytes. Flow cytometry detected twice as many immature neutrophils compared to microscopy. CONCLUSIONS:: We have designed a one-tube immunophenotyping panel for classification of total nucleated cells and platelets in blood or bone marrow. The seven parameters available in one single tube in our cytometer seem to be enough for reliable differential count even in difficult pathological samples. The analytical imprecision of the flow cytometer differential was much lower than that obtained with microscopy or cell counter differentials. (c) 2007 Clinical Cytometry Society.
|
|
| 2. |
|
|
| 3. |
|
|
| 4. |
- Gupta, Monali, et al.
(författare)
-
Radar plots facilitate differential diagnosis of acute promyelocytic leukemia and NPM1+ acute myeloid leukemia by flow cytometry
- 2021
-
Ingår i: Cytometry Part B - Clinical Cytometry. - : Wiley. - 1552-4949 .- 1552-4957. ; 100:4, s. 409-420
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Acute promyelocytic leukemia (APL) is one of the most life-threatening hematological emergencies and requires a prompt correct diagnosis by cytomorphology and flow cytometry (FCM) with later confirmation by cytogenetics/molecular genetics. However, nucleophosmin 1 muted acute myeloid leukemia (NPM1+ AML) can mimic APL, especially the hypogranular variant of APL. Our study aimed to develop a novel, Radar plot-based FCM strategy to distinguish APLs and NPM1+ AMLs quickly and accurately. Method: Diagnostic samples from 52 APL and 32 NPM1+ AMLs patients were analyzed by a 3-tube panel of 10-color FCM. Radar plots combining all markers were constructed for each tube. Percentages of positive leukemic cells and mean fluorescence intensity were calculated for all the markers. Results: APL showed significantly higher expression of CD64, CD2, and CD13, whereas more leukemic cells were positive for CD11b, CD11c, CD15, CD36, and HLA-DR in NPM1+ AMLs. Radar plots featured CD2 expression, a lack of a monocytic component, lack of expression of HLA-DR and CD15, and a lack of a prominent CD11c+ population as recurring characteristics of APL. The presence of blasts with low SSC, presence of at least some monocytes, some expression of HLA-DR and/or CD15, and a prominent CD11c population were recurrent characteristics of NPM1+ AMLs. Radar plot analysis could confidently separate all hypergranular APL cases from any NPM1+ AML and in 90% of cases between variant APL and blastic NPM1+ AML. Conclusion: Radar plots can potentially add to differential diagnostics as they exhibit characteristic patterns distinguishing APL and different types of NPM1+ AMLs.
|
|
| 5. |
- Hammarsten, Ola, et al.
(författare)
-
Use of the cell division assay to diagnose Fanconi anemia patients' hypersensitivity to mitomycin C
- 2021
-
Ingår i: Cytometry Part B-Clinical Cytometry. - : Wiley. - 1552-4949 .- 1552-4957. ; 100:3, s. 1894-1906
-
Tidskriftsartikel (refereegranskat)abstract
- The recently reported cell division assay (CDA) was optimized to measure the relative sensitivity of cells to cytotoxic drugs in vitro. Here, we investigated the in vitro hypersensitivity of lymphocytes from Fanconi anemia (FA) patients, to cytotoxic drugs using CDA. Peripheral blood mononuclear cells (PBMC) as well as cell lines derived from FA patients were treated with two DNA interstrand crosslinking (ICL) agents, mitomycin C and cyclophosphamide. Our data indicate that the CDA detects hypersensitivity of cells from FA patients to mitomycin C. Further, cell lines derived from FA-patients were also hypersensitive to mitomycin C as well as cyclophosphamide, when assayed by the CDA. This study suggests that the CDA is a useful alternative for the diagnosis of FA patients' hypersensitivity to ICL agents.
|
|
| 6. |
- Johansson, Pegah, 1978, et al.
(författare)
-
Validation of a flow cytometry-based detection of γ-H2AX, to measure DNA damage for clinical applications.
- 2017
-
Ingår i: Cytometry. Part B, Clinical cytometry. - : Wiley. - 1552-4957 .- 1552-4949. ; 92, s. 534-540
-
Tidskriftsartikel (refereegranskat)abstract
- The nucleosomal histone protein H2AX is specifically phosphorylated (γ-H2AX) adjacent to DNA double-strand breaks (DSBs) and is used for quantifying DSBs. Many chemotherapies and ionizing radiation (IR) used in cancer treatment result in DSBs. Therefore, γ-H2AX has a significant potential as a biomarker in evaluating patient sensitivity and responsiveness to IR and chemotherapy.
|
|
| 7. |
- Kern, Wolfgang, et al.
(författare)
-
Multicenter prospective evaluation of diagnostic potential of flow cytometric aberrancies in myelodysplastic syndromes by the ELN iMDS flow working group
- 2023
-
Ingår i: Cytometry Part B - Clinical Cytometry. - : Wiley. - 1552-4949 .- 1552-4957. ; 104:1, s. 51-65
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Myelodysplastic syndromes (MDS) represent a diagnostic challenge. This prospective multicenter study was conducted to evaluate pre-defined flow cytometric markers in the diagnostic work-up of MDS and chronic myelomonocytic leukemia (CMML). Methods: Thousand six hundred and eighty-two patients with suspected MDS/CMML were analyzed by both cytomorphology according to WHO 2016 criteria and flow cytometry according to ELN recommendations. Flow cytometric readout was categorized ‘non-MDS’ (i.e. no signs of MDS/CMML and limited signs of MDS/CMML) and ‘in agreement with MDS’ (i.e., in agreement with MDS/CMML). Results: Flow cytometric readout categorized 60% of patients in agreement with MDS, 28% showed limited signs of MDS and 12% had no signs of MDS. In 81% of cases flow cytometric readouts and cytomorphologic diagnosis correlated. For high-risk MDS, the level of concordance was 92%. A total of 17 immunophenotypic aberrancies were found independently related to MDS/CMML in ≥1 of the subgroups of low-risk MDS, high-risk MDS, CMML. A cut-off of ≥3 of these aberrancies resulted in 80% agreement with cytomorphology (20% cases concordantly negative, 60% positive). Moreover, >3% myeloid progenitor cells were significantly associated with MDS (286/293 such cases, 98%). Conclusion: Data from this prospective multicenter study led to recognition of 17 immunophenotypic markers allowing to identify cases ‘in agreement with MDS’. Moreover, data emphasizes the clinical utility of immunophenotyping in MDS diagnostics, given the high concordance between cytomorphology and the flow cytometric readout. Results from the current study challenge the application of the cytomorphologically defined cut-off of 5% blasts for flow cytometry and rather suggest a 3% cut-off for the latter.
|
|
| 8. |
|
|
| 9. |
- Langenskiöld, Cecilia, et al.
(författare)
-
Determination of blood cell subtype concentrations from frozen whole blood samples using TruCount beads
- 2018
-
Ingår i: Cytometry Part B - Clinical Cytometry. - : Wiley. - 1552-4949 .- 1552-4957. ; 94:4, s. 660-666
-
Tidskriftsartikel (refereegranskat)abstract
- © 2016 International Clinical Cytometry Society.Background: In many studies it would be advantageous if blood samples could be collected and analyzed using flow cytometry at a later stage. Ideally, sample collection should involve little hands-on time, allow for long-term storage, and minimally influence the samples. Methods: Here we establish a flow cytometry antibody panel that can be used to determine granulocytes, monocytes, and lymphocyte subset concentrations in fresh and frozen whole blood using TruCount technology. Results: The panel can be used on fresh whole-blood samples as well as whole-blood samples that have been frozen after mixing with 10% DMSO. Concentrations in frozen and fresh sample is highly correlated both when frozen within 4 h and the day after collection (r≥0.98), and the estimated concentration in frozen samples was between 91 and 94% of that in fresh samples for all cell types. Conclusion: Using this method whole-blood samples can be frozen using a simple preparation method, and stored long-term before accurate determination of cell concentration. This allows for standardized analysis of the samples at a reference laboratory in multi-center studies.
|
|
| 10. |
|
|
