All lanes : Anti-Pyruvate Dehydrogenase E2 antibody [15D3G9C11] (ab110332) at 0.5 µg

Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : DLAT knockout HAP1 whole cell lysate
Lane 3 : HeLa whole cell lysate
Lane 4 : HL-60 whole cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 69 kDa

Lanes 1 - 4: Merged signal (red and green). Green - ab110332 observed at 72 kDa. Red - loading control, ab181602, observed at 38 kDa.

ab110332 was shown to specifically react with in wild-type HAP1 cells as signal was lost in DLAT knockout cells. Wild-type and DLAT knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% Milk. Ab110332 and ab181602 (Rabbit anti-GAPDH loading control) were incubated overnight at 4°C at 0.5 μg/ml and 1/20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

Immunocytochemistry analysis using ab110332 at 1µg/ml staining Pyruvate Dehydrogenase E2 in cultured, normal Human embryonic lung fibroblasts (strain MRC5) and an AlexaFluor® 488 goat anti-mouse IgG1 secondary antibody (2 ug/ml).

Immunohistological analysis using ab110332 at 1/100 dilution staining Pyruvate Dehydrogenase E2 in Human cerebellum tissue (Formalin-fixed, Paraffin-embedded).

ICC/IF image of ab109866 stained HepG2 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab109866 at 10µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- mouse (ab96879) IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

Anti-Pyruvate Dehydrogenase E2 antibody [15D3G9C11] (ab110332) at 0.5 µg/ml + Isolated mitochondria from Human heart at 5 µg

Predicted band size: 69 kDa

ICC/IF image of ab109866 stained HepG2 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab109866 at 10µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- mouse (ab96879) IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

Flow cytometric analysis using ab110332 at 1µg/ml staining Pyruvate Dehydrogenase E2 in HL60 cells (blue). Isotype control antibody (red).