Recombinant Anti-IDH2 antibody [EPR7577] - BSA and Azide free (ab230796)

This WB data was generated using the same anti-IDH2 antibody clone, EPR7577, in a different buffer formulation (cat# ab131263).

Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: IDH2 knockout HAP1 cell lysate (20 µg) 
Lane 3: K562 cell lysate (20 µg)
Lane 4: HepG2 cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab131263 observed at 48 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab131263 was shown to recognize IDH2 when IDH2 knockout samples were used, along with additional cross-reactive bands. Wild-type and IDH2 knockout samples were subjected to SDS-PAGE. ab131263 and ab8245 (loading control to GAPDH) were both diluted 1/10 000 and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

Overlay histogram showing MCF7 cells stained with ab131263 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab131263, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1µg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in MCF7 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab131263).

Equilibrium disassociation constant (K_D)
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab131263).

This IHC data was generated using the same anti-IDH2 antibody clone, EPR7577, in a different buffer formulation (cat# ab131263).

Immunohistochemical analysis of paraffin embedded Human liver tissue labelling IDH2 with ab131263 at 1/100 dilution.

Heat mediated antigen retrieval was performed with citrate buffer pH 6 before commencing with IHC staining protocol.