Establishment of the human PBMC-engrafted mouse model

This study was approved by the Institutional Review Board of Peking University People’s Hospital. Blood samples were collected from donors with written informed consent. NPG mice (NOD prkdc^scidIl2rg^null) were purchased from Beijing Vitalstar Biotech. Co. Ltd. The two- to threefold diluted blood samples were subjected to centrifugation on a lymphocyte separation medium (Tianjin Haoyang Biological Manufacture Co. Ltd.) at a density of 1.077 g/ml, and the nucleated cell layer between the plasma and separation medium was gathered. After two washes with RPMI 1640 medium, the PBMC pellets were suspended in RPMI 1640 medium at a density of 5 × 10⁷ cells/ml. Then, 1 × 10⁷ cells were injected via tail vein for each NPG mouse. Mice were housed in an SPF facility and accessed food and water ad libitum. The PBMC-transplanted mice were bled retro-orbitally every week, and the human CD45-positive cell rate in the mouse peripheral blood was analyzed by flow cytometry. The mice with more than 25% human CD45-positive cells in their blood were considered successful human PBMC-engrafted mouse models (see Additional files 1 and 2) and subjected to tumor cell inoculation.