Anti-Human CCR4 Recombinant Antibody (406)

ScFvs were cross-linked using anti-Myc-antibody (mouse) to simulate a dimeric situation and diluted 3-fold, starting at 10 μg/mL. Bound scFvs were detected using RPE-conjugated anti-mouse IgG. a.) Binding to CCR4-positive DT40 cells, b.) Binding to CCR4-negative DT40 cells.

ScFvs were cross-linked using anti-Myc-antibody (mouse) to simulate a dimeric situation and diluted 3-fold, starting at 20 μg/mL. Bound scFvs were detected using RPE-conjugated anti-mouse IgG.

ScFvs were incubated at a concentration of 5 μg/mL (150 nM) and diluted 3-fold over twelve dilution points in the presence of a fixed concentration of MDC-SNAP (50 nM).

ScFvs were incubated at a concentration of 0.5 μg/mL in presence of 2.5 μg/mL mouse anti-Myc-antibody (mouse) in presence of a fixed concentration of ligands (1 μg/mL) and stained using an anti-mouse-RPE conjugated antibody. Signals were compared to cells, stained only with antibody-controls (background) as well as to unstained cells (unstained).

The biotinylated KW0761-lgG was detected using PE-conjugated Streptavidin.

TARC was incubated at a final concentration of 28.6 ng/μL (3.6 nM) in presence of increasing scFv-concentrations, titrated over six dilution points, starting at 0.5 μg/mL. The signals were expressed in % where 100% signaling refers to recorded signals in presence of TARC-ligand, but no scFv antibody.

MDC was incubated at a final concentration of 5 ng/μL (6.25 nM) in presence of a fixed scFv concentration of 10 μg/mL. The signals were expressed in % where 100% signaling refers to recorded signals in presence of MDC-ligand, but no scFv antibody.

TARC had a fixed concentration of 3.5 nM. The signals were expressed in % where 100% refers to migration of cells in presence of TARC-ligand, but no scFv antibody. The graphs were fitted.

IgGs were incubated at defined concentrations and bound IgGs were detected using RPE-conjugated anti-Human IgG. a.) Binding to CCR4-positive CCRF-CEM cells. IgGs were diluted four-fold over eight points starting at 1 μg/ml. b.) Binding to CCR4-positive Hut78 cells. IgGs were diluted four-fold over eight points starting at 1 μg/ml. c.) Binding to CCR4-positive L-428 cells. IgGs were diluted three-fold over eight points starting at 10 μg/ml.

Biotinylated samples were incubated at 10 μg/ml in either 0% human serum (FACS-buffer) or in 10 and 50% human serum. Samples were detected via PE-conjugated-Streptavidin. a.) Comparison of recorded binding signals, b.) Remaining binding signals converted into %.