Percp cy5 5 conjugated anti mcd45

Peripheral blood was obtained at the time of first AraC dose (day 0) and at the time of dissection (day 8) two days after the last dose of AraC to determine the fraction of human blasts using flow cytometry. NSG mice were humanely killed in accordance with European ethic protocols. Bone marrow (mixed from tibias and femurs) and spleen were dissected in a sterile environment and flushed in Hanks balanced salt solution with 1% FBS. MNCs from peripheral blood, bone marrow and spleen were labeled with FITC-conjugated anti-hCD3, PE-conjugated anti-hCD33, PerCP-Cy5.5-conjugated anti-mCD45.1, APC-conjugated anti-hCD45 and PeCy7-conjugated anti-hCD44 (all antibodies from Becton Dickinson, BD, except FITC-conjugated anti-hCD3 from Ozyme Biolegend) to determine the fraction of human blasts (hCD45⁺mCD45.1⁻hCD33⁺hCD44⁺ cells) using flow cytometry. Analyses were performed on a Life Science Research II (LSR II) flow cytometer with DIVA software (BD) or Cytoflex flow cytometer with CytoExpert software (Beckman Coulter). The number of AML cells/ul peripheral blood and number of AML cells in total cell tumor burden (in bone marrow and spleen) were determined by using CountBright beads (Invitrogen) using described protocol.