Lymphocytes were defined in a graph of size and complexity (A) (FSC-A x SSC-A) and analyzed as singlets (B) (SSC-W x SSC-H). After excluding CD3+ cells and selecting CD19+ (C) (CD3 x CD19) and CD20+ cells (D) (CD20 x SSC), B cell subpopulations were defined (E) (CD43 x CD27) as naïve (CD43- CD27-), memory (CD43- CD27+) and B1 cells (CD43+ CD27+). This scheme is representative of all analyzed samples. About 2,000,000 total events were acquired, and the entire sample was submitted to cell sorting.