Anti-Human IL-6 Recombinant Antibody (Olokizumab) (CAT#: TAB-253)

Figure 1 Olokizumab in cell assays to assess neutralization of IL-6 activity.

(A) Cis-signaling: Primary human hepatocytes were cultured in collagen-coated plates and stimulated with IL-6 (12.5 ng/mL) in the presence or absence of olokizumab (titration from 10 µg/mL) for 72 h. Cell supernatants were analyzed for acute phase proteins CrP and SAA using a Luminex kit. Data are plotted with standard errors of means from triplicate determinations in three experiments. (B) Trans-signaling: Different concentrations of olokizumab were pre-incubated with IL-6 at 25ng/mL followed by addition of soluble IL-6 receptor gp80 at 125 ng/mL. This complex was added to prepared HUVeC cells and incubated at 37 °C for 20 min to allow IL-6-induced STAT-3 phosphorylation to occur. The activation was stopped by the addition of ice-cold lysis buffer, and cell supernatants were analyzed for STAT3 phosphorylation using a MSD STAT-3 kit. Data are plotted with standard errors of means from triplicate determinations in two experiments.

Shaw, S. , Bourne, T. , Meier, C. , Carrington, B. , Gelinas, R. , & Henry, A. , et al. (2014). Discovery and characterization of olokizumab. mAbs, 6(3), 773-781.

Figure 1 Anti-Human IL6 Recombinant Antibody (TAB-253) in ELISA

ELISA analysis of TAB-253 was performed by coating with human IL6 protein (His tag). Then blocked with BSA and incubated with TAB-253. The HRP-conjugated goat anti-IgG was used as a secondary antibody (1: 2000). Detection was performed using TMB substrate and stopped with sulfuric acid. The absorbances were read on a spectrophotometer at 450 nm.

Figure 2 Anti-Human IL6 Recombinant Antibody (TAB-253) in WB

Western blot analysis of TAB-253 was performed by loading human IL6 protein onto a 12% Tris-HCl polyacrylamide gel. Proteins were transferred to a CN membrane and blocked with 5% skim milk for at least one hour. Membranes were probed with TAB-253 and a HRP goat anti-IgG (H+L) was used as the secondary antibody (1: 3000). Chemiluminescent detection was performed.

Figure 3 Anti-Human IL6 Recombinant Antibody (TAB-253) in DB

Dot Blot analysis of TAB-253 was performed by coating with human IL6 protein (His tag).
TAB-253 incubation concentration: 2 μg/mL.
HRP-conjugated goat anti-IgG (H+L) secondary antibody (1: 3000)
NC: Negative control, PC: Positive control

Figure 4 Anti-Human IL-6 Recombinant Antibody (TAB-253) in SDS-PAGE

SDS-PAGE analysis of TAB-253 in non-reduced (Lane 1) and reduced (Lane 2) conditions.

Figure 5 Anti-Human IL-6 Recombinant Antibody (TAB-253) in HPLC

HPLC analysis of TAB-253 shows the purity is>95%.