Anti-ATP citrate lyase antibody [EP704Y] (ab40793)

  Western blot - ATP citrate lyase antibody [EP704Y] (ab40793)

Anti-ATP citrate lyase antibody [EP704Y] (ab40793) at 1/5000 dilution + Hela cell lysate.

Predicted band size : 122 kDa
Observed band size : 122 kDa

  Immunocytochemistry/ Immunofluorescence-ATP citrate lyase antibody [EP704Y](ab40793)

ICC/IF image of ab40793 stained MCF7 cells. The cells were 4% PFA 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 (ab40793, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 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 Cytometry - ATP citrate lyase antibody [EP704Y] (ab40793)

Overlay histogram showing HeLa cells stained with ab40793 (red line). The cells were fixed with 80% methanol (5 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 (ab40793, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1μg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.

  Immunoprecipitation - Anti-ATP citrate lyase antibody [EP704Y] (ab40793)

ATP citrate lyase was immunoprecipitated using 0.5mg Hela whole cell extract, 10ug of Rabbit monoclonal [EP704Y] to ATP citrate lyaseand 50µl of protein G magnetic beads (lane 1). The antibody was incubated with the Protein G beads for 10min under agitation. No antibody was added to the control (lane 2). Hela whole cell extractdiluted in RIPA buffer was added to each sample and incubated for 10min under agitation. Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70^oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab40793. Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697). Bands: 122kDa: ATP citrate lyase; non specific - 60kDa: We are unsure as to the identity of this extra band.