Specific killing of aggressive lymphoma cells using BCR-antigen/toxin constructs. (A) LDH-release assay. Percentage specific lysis of nontransfected OCI-Ly3 cells or OCI-LY3 cells transfected to express either MAZ-specific or SAM-specific BCRs after incubation with 5 μg/mL neurabin-I/ETA or MAZ/ETA, respectively, for 24 hours. Experiments were performed in triplicates and repeated 3 times (****P ≤ .0001). (B) Trypan blue exclusion assay. Cell viability of transfected OCI-LY3 cells expressing SAMD14/neurabin-I-specific BCRs or nontransfected OCI-Ly3 cells after incubation with 2 μg/mL neurabin-I/ETA or MAZ/ETA was determined by trypan blue staining after 24 and 48 hours, showing the time-course effect. (C) Dose dependency of cytotoxicity determined by LDH release assay. Percentage specific lysis of transfected TMD8 cells either doxycycline-induced or noninduced after incubation with different concentrations (1.25-10 μg/mL neurabin-I/ETA′ or LRPAP1/ETA′ immunotoxins for 24 hours), showing a dose-dependent effect. (D-E) Annexin-V assay. Characterization of transfected TMD8 and U2932 cells with either doxycycline-induced expression of SAMD14/neurabin-I reactive BCR or noninduced, nonexpressing recombinant BCRs by annexin-V and PI staining after 24-hour cultivation in the presence of SAM/ETA, LRPAP1/ETA, or Staurosporine. In the bottom line, bar graphs represent the proportion of vital, early, late, and total apoptotic cells.