Panel A: DFO-ATZ conjugate. Representative deconvolution results from LC-MS analysis (n = 3). Plot represents counts (y-axis) vs. de-convoluted mass (x-axis). N is the chelator number per ATZ; R0, R1, R2 or Rn is the percentage of the peak area corresponding to DFO-ATZ conjugates with 0, 1, 2, or n DFO chelators, respectively. Panel B: Flow cytometry cell binding assay in A20 cells expressing PD-L1. Data are presented as mean ± standard deviation (s.d.); one representative experiment shown (n = 3 in triplicates); MFI: Mean fluorescence intensity; EC50 values presented as mean ± standard error of means. Panel C: Immunoreactivity assay of [89Zr]Zr-DFO-ATZ in MDA-MB-231 cells expressing PD-L1. Data are presented as mean ± s.d.; one representative experiment shown (n = 6 in triplicates). Panel D: Blocking cell assay to test binding specificity of [89Zr]Zr-DFO-ATZ. One hundred times excess unlabeled ATZ (~ 6 ng) was added to the cell suspension along with the [89Zr]Zr-DFO-ATZ (n = 3). Data are presented as mean ± s.d.