Binding to cell surface receptors
murine myeloma NS0 and Daudi, NS0-PD-1FACS, NS0/PD-1
SECONDARY
Generation of murine myeloma NS0 and Daudi cell lines expressing human PD-1 and LAG-3. NS0/PD-1 was established by stable transfection of the parental NS0 cell line with the Fluorescence-activated cell sorting (FACS) analysis of tebotelimab or nivolumab replica or relatlimab replica binding to NS0 cells transfected with human PD-1 or LAG-3. NS0/PD-1 or NS0/LAG-3 cells were suspended in FACS blocking buffer (FACS buffer with 10% human AB serum) and incubated with testing molecules at the indicated concentrations at 4–25 °C for 30–120 min. Cells were washed and resuspended in FACS blocking buffer. For secondary staining, a goat anti-human Fc-APC (Jackson ImmunoResearch) was used to detect human primary antibodies. The stained cells were acquired on a BD FACSCalibur or Fortessa, and data were analyzed using FlowJo software.FACS analysis of soluble PD-L1 or PD-L2 binding to NS0-PD-1FACS analysis of soluble LAG-3 binding to major histocompatibility complex (MHC)-II-expressing Daudi cells in the presence of titrating concentrations of mAbs or tebotelimab. Daudi cells were suspended in FACS blocking buffer and incubated with 0.5 μg ml