IMPA1 Antibody / Inositol monophosphatase 1 (FY13287)

Immunoprecipitating IMPA1 in Jurkat whole cell lysate. Western blot analysis of IMPA1 using anti-IMPA1 antibody. Lane 1: Jurkat whole cell lysates (30ug), Lane 2: Rabbit control IgG instead of anti-IMPA1 antibody in Jurkat whole cell lysate, Lane 3: anti-IMPA1 antibody (2ug) + Jurkat whole cell lysate (500ug). After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-IMPA1 antibody at a dilution of 0.5 ug/ml and probed with a goat anti-rabbit IgG-HRP secondary antibody. The signal is developed using ECL Plus Western Blotting Substrate. A specific band was detected for IMPA1 at approximately 30 kDa. The expected molecular weight of IMPA1 is ~30 kDa.

Western blot analysis of IMPA1 using anti-IMPA1 antibody. Electrophoresis was performed on a 10% SDS-PAGE gel at 80V (Stacking gel) / 120V (Resolving gel) for 2 hours. Lane 1: human Jurkat whole cell lysates, Lane 2: rat kidney tissue lysates, Lane 3: rat brain tissue lysates, Lane 4: mouse kidney tissue lysates, Lane 5: mouse brain tissue lysates, Lane 6: mouse NIH/3T3 whole cell lysates. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-IMPA1 antibody at 0.5 ug/ml overnight at 4oC, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal was developed using an ECL Plus Western Blotting Substrate. A predominant band is detected at an approximately 30 kDa in all samples, consistent with the predicted size of monomeric IMPA1. Most samples also show a stronger band near 85 kDa, which likely represents an SDS resistant oligomeric or complex form of IMPA1 rather than a distinct isoform.

Flow Cytometry analysis of Jurkat cells using anti-IMPA1 antibody. Overlay histogram showing Jurkat cells stained with (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-IMPA1 antibody (1 ug/million cells) for 30 min at 20oC. DyLight 488 conjugated goat anti-rabbit IgG (5-10 ug/million cells) was used as secondary antibody for 30 minutes at 20oC. Isotype control antibody (Green line) was rabbit IgG (1 ug/million cells) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.