Background: Monoclonal B lymphocytosis (MBL) increases with age and individuals with high count MBL progress to chronic lymphocytic leukaemia requiring therapy at a rate of ~1%-5% per year. These cases usually have atypical lymphocytes at the microscope, abnormal representation in the scattergram, and positivity of flags. Using XN9000 (Sysmex), we noticed cases of MBL without this correlation. We studied customized gates for discovering MBL cases of our interest. Methods: We considered 212 peripheral blood samples with known phenotypes: 76.7% negative and 23.3% positive for B, T, or NK lymphocytes clones. We created gates studying the XN9000 FCS files in Diva software to identify new areas for better delimiting subpopulations of our interest and calculating sensitivity and specificity. Results: We found significant differences between negative and positive groups for Q-flag "Blasts/Abn Lympho?" (B/AL) and LY-X (p <0.05) with lymphocyte counts below 5×109/L. A new gate P1 normalized by P2 (P1n) differentiated between phenotypes much better than Q-flag B/AL with lymphocyte counts ≤ 5×109/L. Moreover, cases with MBL CD5 positive had higher medians (p <0.05). Conclusion: We propose a gate P1n as a new Q-flag for lymphocytes count ≤ 5×109/L, in order to hypothesize the presence of MBL CD5 positives.
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