Alexa Fluor® 488 Anti-ATP citrate lyase antibody [EP704Y]

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Alexa Fluor® 488 Anti-ATP citrate lyase antibody [EP704Y] (AB205429)

Flow cytometry overlay histogram showing left wild-type HAP1 positive cells and right negative ACLY knockout HAP1 stained with ab205429 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab205429) (1x 10⁶ in 100μl at 0.04 μg/ml (1/12500)) for 30min at 22°C.

Isotype control antibody was Alexa Fluor^® 488 Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab199091) (black line) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

This antibody gave a positive signal in wild-type HAP1 fixed with 4% formaldehyde (10 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min under the same conditions.

This data was acquired using a Beckman Coulter CytoFlexLX flow cytometer.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Alexa Fluor® 488 Anti-ATP citrate lyase antibody [EP704Y] (AB205429)

Flow cytometry scatter plots showing human peripheral mononuclear cells (PBMCs) stained with ab205429 (right) or isotype control (left). The cells were fixed and permeabilised using BD CytoFix/CytoPerm™ (20 min). The cells were incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab205429) (1x 10⁶ in 100μl at 0.2 μg/ml (1/2500)) for 30min on ice. Alexa Fluor^® 488 Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab199091) (left). The cells were simultaneously stained with CD3.

Acquisition of >30000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

This data was acquired using a Beckman Coulter CytoFlexLX flow cytometer.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Alexa Fluor® 488 Anti-ATP citrate lyase antibody [EP704Y] (AB205429)

ab205429 staining ATP citrate lyase in HeLa cells. The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab205429 at 1/500 dilution (shown in green) and ab195889, Mouse monoclonal [DM1A] to alpha Tubulin (Alexa Fluor^® 594), at 2μg/ml (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).