Chimeric antigen receptor (CAR) T cell therapy is an innovative immunotherapy for treating cancers in both children and adults with proven utility in numerous clinical trials. Significantly, some CAR T cell therapies have now been approved by relevant national regulatory bodies across numerous countries for clinical therapeutic use outside of clinical trials. One such recently licensed product is tisagenlecleucel, a CAR T therapy approved for the treatment of B-cell acute lymphoblastic leukemia (B-ALL) using autologous T cells from the patient. The genetically engineered T cells target a protein called CD19, common to B cells, through a CAR incorporating a 4-1BB costimulatory domain to improve response. Since tisagenlecleucel is now a standard of care treatment for B-ALL, it is clinically essential to be able to accurately monitor these CAR T cells in patients. Assessment of the copy number variant (CNV) of the CAR T cell products allows this within a clinically acceptable timeframe for optimal patient benefit. However, no standardized method with high reproducibility and efficiency has been described within a routine clinical laboratory setting. Here, we demonstrated a novel digital droplet PCR (ddPCR)-based methodology for the study of CNV (ddPCR-CNV) in 4-1BB CD19-specific CAR T cells with universal applicability across clinical diagnostic laboratories.