NeuroMab™ Anti-CD54 Antibody(NRP-0422-P1406)

Figure 1 Binding of seven unique scFv clones to Ramos cells (solid bars) and Jurkat cells (open bars). Control scFv (ctrl) did not bind to any cells.

Figure 2 Anti-Ramos scFv induces apoptosis.

Ramos B lymphoma cells were incubated sequentially with anti-Ramos scFv, anti-His mAb, and anti-mouse polyclonal antibody on ice (intermittent washing to remove excess unbound antibody) and incubated at 37 °C in a humidified atmosphere with 5% CO2 C for 24 hours. Cells were then harvested and co-stained with AnnexinTM V-AF488 (AV) and propidium iodide (PI).

Figure 3 Anti-Ramos scFv induces apoptosis.

Seven unique scFv clones were incubated with Ramos B lymphoma cells at different concentrations for 24 hours at 37°C, and the effect on apoptosis induction was studied. Three scFvs; Bl, B11 and C11 showed titratable activity against both cell lines, whereas apoptosis-inducing

Figure 4 Specificity of isolated antibodies including HLA-DR/DP, IgM and ICAM-1.

50-600 x 10^6 Raji B lymphoma cells were lysed with 0.5% v/v of the non-ionic detergent Triton X100TM and immunized with 100 μg of fully human IgG1 versions of B1 (lane 1) and B11 (lane 2) Antibody is precipitated and then cross-linked with Protein A SepharoseTM. Ramos B lymphoma cell lysate from 50 x 106 cells was used to pellet 20 μg C11 (lane 3).

Figure 5 Specificity of isolated antibodies including HLA-DR/DP, IgM and ICAM-1.

Binding of B1 IgG, B11 IgG and Cll IgG to B lymphoma cells was specifically blocked by preincubation with anti-HLA-DR/DP, anti-ICAM-1 or anti-IgM antibodies, respectively.

Figure 8 Binding of B1, B11, C11 IgG1 to tumor cell lines of different origins. The antigen distribution of the antigens targeted by the B1, B11 and C11 antibodies on different cancer cell lines was studied by flow cytometry.

Figure 9 Induction of apoptosis in B11 IgG1 cells in cancer cells.

Bar graphs show the mean percentage of apoptotic PC-3 cells after incubation with serially diluted B11 IgG1 or 20 g/m1 B1 IgG1.

Figure 10 B1 IgG1 and B1 IgG4 induced direct cytotoxicity against Raji B lymphoma cells in the absence of crosslinker.

Raji cells were incubated with B1 IgG1, B1 IgG4, rituximab IgG1 or control CT-17 IgG1 at 20, 6.7 or 2.2 μg/ml for 24 hours. Cells were harvested and viability was determined as a percentage of Annexin V and propidium iodide double negative cells.