A 5-Color Flow Cytometric Method for Extended 8-Part Leukocyte Differential

Objectives: Microscopic leukocyte differentials display many drawbacks. Several single 5 to 8-color tubes using multiparameter flow cytometry (MFC) are able to provide extended differentials with sequential gating-based analysis strategies. We investigated a new 5-color MFC method to perform an extended 8-part differential with a simplified gating strategy. Methods: Whole blood was stained with a combination of antibodies including HLA-DR-FITC/CD19-PE/CD45-ECD/CD16-PC5+CD71-PC5/CD5-PC7. Results: An original approach was developed to exclude debris and straightforwardly gate the cells to identify sixteen populations. Strong correlations were obtained with the analyzer for neutrophils, lymphocytes, monocytes, and eosinophils (R-2>0.9). Abnormal cells, such as immature granulocytes and blast cells were identified with a good sensitivity and excellent correlation against cytomorphologic review (R-2=0.66 and 0.99, respectively). The choice of HLA-DR and CD5 improved specificity for the identification of activated T-lymphocytes and some lymphoid neoplastic cells, respectively. Conclusions: Here a new cytometric differential is proposed with a robust gating strategy which may be used even by unskilled cytometrists and can be easily automated. (C) VC 2017 International Clinical Cytometry Society