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Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human PI 3 Kinase catalytic subunit alpha aa 1000 to the C-terminus (C terminal). The exact sequence is proprietary.
A trial size is available to purchase for this antibody.
Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.
This product is a recombinant rabbit monoclonal antibody.
Our Abpromise guarantee covers the use of ab40776 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use at an assay dependent concentration.|
|WB||1/1000 - 1/10000. Detects a band of approximately 110 kDa (predicted molecular weight: 110 kDa).|
|ICC/IF||1/100 - 1/250.|
|IP||1/20 - 1/30.|
ab40776 staining PI 3 Kinase catalytic subunit alpha in the human cell line Jurkat (human acute T cell leukemia) by flow cytometry. Cells were fixed with 4% paraformaldehyde, permiabilised with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/40. A goat anti rabbit IgG (Alexa Fluor® 488) at a dilution of 1/2000 was used as the secondary antibody.
Isoytype control: Rabbit monoclonal IgG (Black)
Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)
Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: PI 3 Kinase catalytic subunit alpha knockout HAP1 cell lysate (20 µg)
Lane 3: Jurkat cell lysate (20 µg)
Lane 4: MCF7 cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab40776 observed at 120 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab40776 was shown to recognize PI 3 Kinase catalytic subunit alpha when PI 3 Kinase catalytic subunit alpha knockout samples were used, along with additional cross-reactive bands. Wild-type and PI 3 Kinase catalytic subunit alpha knockout samples were subjected to SDS-PAGE. ab40776 and ab8245 (loading control to GAPDH) were diluted to 1/1000 and 1/10000 respectively 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 hour at room temperature before imaging.
Lanes 1 - 3: Merged signal (red and green). Green - ab40776 observed at 120 kDa. Red - loading control, ab8245, observed at 37 kDa.
This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab40776 and ab8245 (loading control) overnight at 4°C. Antibody binding was detected using Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at a 1:10000 dilution for 1hr at room temperature and then imaged.
Immunocytochemistry/Immunofluorescence analysis of Jurkat cells labelling PI 3 Kinase catalytic subunit alpha with purified ab40776 at a dilution of 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.
Control 1: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000).
ab40776 (purified) at a dilution of 1/20 immunoprecipitating PI 3 Kinase catalytic subunit alpha in Jurkat whole cell lysate.
Lane 1 (input): Jurkat whole cell lysate (10µg)
Lane 2 (+): ab40776 + Jurkat whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab40776 in Jurkat whole cell lysate.
For western blotting, ab131366 VeriBlot for IP (HRP) was used as the secondary antibody (1/1000).
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling PI 3 Kinase catalytic subunit alpha with unpurified ab40776 at a dilution of 1/100.
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"