PDF
ePub
Citation
Print
Abstract
Background
The lineage assignment in acute leukemias is critical in therapeutic decisions. Immunophenotyping by flow cytometry plays the main role in the lineage assignment; however, few studies have been done to determine the optimal set of markers. In this regard, we tried to find out the optimal first-line set of markers with a minimal compromise in its diagnostic sensitivity.
Materials and Methods
We retrospectively analyzed 321 cases of acute leukemias whose diagnoses were based on the EGIL (European Group for Immunological Classification of Acute Leukemia) scores. At our institution, flow cytometic analyses included 15 first-line markers and 4 additional second-line markers as needed, along with immunohistochemical stains. We performed simulational studies for the expected EGIL scores involving every possible combination of markers and analyzing the overall diagnostic sensitivities in each combination.
Results
The cytoplasmic antigens including MPO stain and CD79a stain contributed greatly to the lineage assignment. For a sensitivity over 95%, there needed a combination of MPO stain with other 5 flow markers (CD33, CD13, CD14, CD15 and CD117) for myeloid lineage; CD79a stain with 3 flow markers [CD19, CD10, and CD20 (or TdT)] for B-lymphoid lineage; and 4 flow markers (CD2, CD3, CD5, and CD7) for T-lymphoid lineage.
Conclusions
To maintain diagnostic sensitivities over 95% for each lineage, at least 14 markers (including MPO stain and CD79a stain) were needed; while 16 markers were needed for a sensitivity of 100%. When combined with other important markers for specific aims such as CD45, the minimum number of markers needed for the accurate diagnosis of acute leukemias would be more than about 18 to 20.
References
1. McCulloch EA. Stem cells in normal and leukemic hemopoiesis (Henry Stratton Lecture, 1982). Blood. 1983; 62:1–13.
2. Bennett JM, Catovsky D, Daniel MT, Flandrin G, Galton DA, Gralnick HR, et al. Proposals for the classification of the acute leukaemias. French-American-British (FAB) co-operative group. Br J Haematol. 1976; 33:451–8.
3. Harris NL, Jaffe ES, Diebold J, Flandrin G, Muller-Hermelink HK, Vardiman J, et al. World Health Organization classification of neoplastic diseases of the hematopoietic and lymphoid tissues: report of the Clinical Advisory Committee meeting-Airlie House, Virginia, November 1997. J Clin Oncol. 1999; 17:3835–49.
4. Tallman MS, Gilliland DG, Rowe JM. Drug therapy for acute myeloid leukemia. Blood. 2005; 106:1154–63.
5. Pui CH, Evans WE. Treatment of acute lymphoblastic leukemia. N Engl J Med. 2006; 354:166–78.
6. Bene MC, Castoldi G, Knapp W, Ludwig WD, Matutes E, Orfao A, et al. Proposals for the immunological classification of acute leukemias. European Group for the Immunological Characterization of Leukemias (EGIL). Leukemia. 1995; 9:1783–6.
7. The value of c-kit in the diagnosis of biphenotypic acute leukemia. EGIL (European Group for the Immunological Classification of Leukaemias). Leukemia. 1998; 12:2038.
8. Jennings CD, Foon KA. Recent advances in flow cytometry: application to the diagnosis of hematologic malignancy. Blood. 1997; 90:2863–92.
9. Hallden G, Andersson U, Hed J, Johansson SG. A new membrane permeabilization method for the detection of intracellular antigens by flow cytometry. J Immunol Methods. 1989; 124:103–9.
10. Braylan RC, Orfao A, Borowitz MJ, Davis BH. Optimal number of reagents required to evaluate hematolymphoid neoplasias: results of an international consensus meeting. Cytometry. 2001; 46:23–7.
11. Bene MC. Immunophenotyping of acute leukaemias. Immunol Lett. 2005; 98:9–21.
12. Basso G, Buldini B, De Zen L, Orfao A. New methodologic approaches for immunophenotyping acute leukemias. Haematologica. 2001; 86:675–92.
13. Lacombe F, Durrieu F, Briais A, Dumain P, Belloc F, Bascans E, et al. Flow cytometry CD45 gating for immunophenotyping of acute myeloid leukemia. Leukemia. 1997; 11:1878–86.
14. Orfao A, Schmitz G, Brando B, Ruiz-Arguelles A, Basso G, Braylan R, et al. Clinically useful information provided by the flow cytometric immunophenotyping of hematological malignancies: current status and future directions. Clin Chem. 1999; 45:1708–17.
Table 1.
The distribution of scores for each lineage according to the types of leukemias
| Lineage | Score | AML | B-ALL | T-ALL | M/B-BAL |
|---|---|---|---|---|---|
| M score | 0 | − | 32/87 | 6/14 | − |
| (36.8%) | (42.9%) | ||||
| 0.5–2 | − | 55/87 | 8/14 | − | |
| (63.2%) | (57.1%) | ||||
| 2.5–4 | 119/187 | − | − | 13/14 | |
| (66.3%) | (92.9%) | ||||
| >4 | 63/187 | − | − | 1/14 | |
| (33.7%) | (7.1%) | ||||
| B score | 0 | 128/187 | − | 9/14 | − |
| (68.4%) | (64.3%)* | ||||
| 0.5–2 | 59/187 | − | 5/14 | − | |
| (31.5%) | (35.7%)* | ||||
| 2.5–4 | − | 34/87 | − | 10/14 | |
| (39.1%) | (71.4%) | ||||
| >4 | − | 53/87 | − | 4/14 | |
| (60.9%) | (28.6%) | ||||
| T score | 0 | 147/187 | 80/87 | − | 14/14 |
| (78.6%)* | (92.0%)* | (100%)* | |||
| 0.5–2 | 40/187 | 7/87 | − | 0 | |
| (21.4%)* | (8.0%)* | ||||
| 2.5∼4 | − | − | 3/14 | − | |
| (21.4%) | |||||
| >4 | − | − | 11/14 | − | |
| (78.6%) |
Table 2.
The concordance rate between flow cytometric markers and immunocytochemical stains
| +/+ | +/- | −/+ | −/- | NC | Concordance rate | |
|---|---|---|---|---|---|---|
| MPO/stMPO | 107 | 2 | 23 | 82 | 101 | 88.3% |
| CD3/stCD3 | 3 | 3 | 2 | 21 | 286 | 82.8% |
| CD20/stCD20 | 35 | 8 | 3 | 45 | 224 | 87.9% |
| TdT/stTdT | 81 | 2 | 2 | 12 | 218 | 95.9% |
| CD14/ANBE | 11 | 4 | 19 | 111 | 170 | 84.1% |
Table 3.
The best combinations and sensitivities for myeloid markers
Table 4.
The best combinations and sensitivities for B-lymphoid markers
Table 5.
The best combinations and sensitivities for T-lymphoid markers
XML Download