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Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human beta Actin.
(Peptide available as ab13772.)
The concentration of this antibody is batch dependent.
Alternative versions available:
Our Abpromise guarantee covers the use of ab8226 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use a concentration of 5 µg/ml.|
|ICC||Use at an assay dependent concentration.|
|Flow Cyt||Use at an assay dependent concentration. Ab170190-Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.|
|IHC-FrFl||Use at an assay dependent concentration.|
|IHC-P||Use at an assay dependent concentration.|
|WB||1/500 - 1/10000. Predicted molecular weight: 42 kDa.Can be blocked with Human beta Actin peptide (ab13772). Abcam recommends blocking and incubation using 5% BSA as we have found that use of 5% milk significantly reduces the intensity of the signal obtained. Please refer to WB image for further details.|
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 5% Milk before being incubated with ab8226 overnight at 4°C. Antibody binding was detected using a goat anti-mouse Alexa 790 (ab175783) at a 1:10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.
Immunofluorescence using ab8226 at 5ug/ml incubated for 1 hour on Rat Colon Cancer cells.
Cells were fixed with ice-cold methanol for 5 mins, then for all following steps, permeabilised in TBS-T for 30 mins, blocked with 5% BSA for 30 mins and then washed in TBS-T. Secondary antibody was Alexa Fluor 488 goat anti-mouse IgG at 1/1000 incubated for 1 hour. Cells were counterstained with DAPI. Image at 400X magnification. All incubations were at room temperature.
The beta actin fibres can be seen arrayed around the edge of the cells.
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 2% Bovine Serum Albumin before being incubated with ab8226 overnight at 4°C. Antibody binding was detected using an anti-mouse HRP (ab97040) antibody conjugated to HRP, and visualised using ECL development solution ab133406
ab8226 staining beta Actin in human AGS cells by Immunocytochemistry. Cells were fixed with formaldehyde, permeabilized with 0.025% Triton-X in TBS and blocking with 5% serum was performed for 1 hour at 230C. Samples were incubated with primary antibody (1/500) for 1 hour at 23°C. An HRP®-conjugated goat polyclonal to mouse IgG was used undiluted as secondary antibody.
ab8226 staining beta Actin in human gastric epithelial AGS cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed in formaldehyde. Permeabilization and blocking was carried out using 5% BSA containing 0.025% Triton X in TBS for 1 hour at 23°C. Primary antibody was used at 5µg/ml for 1 hour at 23°C. An Alexa Fluor 488® conjugated goat polyclonal to mouse IgG was used as the secondary antibody at a 1/1000 dilution.
Western blot image using the Optiblot Reducing Electrophoresis Kit - 10 x 10 cm (4-20%) (ab119220) with the
Prism Ultra Protein Ladder (ab116028) 5µl used. We recommend using our ECL substrate kit (ab65623).
Loading Control (ab8226) used at 1 µg/ml.
Lane 1 : Hela cell lysate
Lane 2 : Jurkat cell lysate
Lane 3 : A431 cell lysate
Lane 4 : HEK293 cell lysate
Lane 5 : HepG2 cell lysate
Lysates loaded at 20 µg per lane.
Lane 1 : beta Actin antibody [mAbcam 8226] - Loading Control (ab8226) at 1/1000 dilution
Lane 2 : beta Actin antibody [mAbcam 8226] - Loading Control (ab8226) at 1/10000 dilution
Lanes 3 - 4 : beta Actin antibody [mAbcam 8226] - Loading Control (ab8226) at 1/500 dilution
Lane 1 : HeLa Cell lysate at 20 ug
Lane 2 : HeLa Cell lysate at 20 ug
Lane 3 : 293 Cell lysate at 20 ug
Lane 4 : 3T3 Mouse Cell lysate at 20 ug
Rabbit polyclonal to Mouse IgG H&L (HRP) (ab6728) at 1/5000 dilution
Performed under reducing conditions.
Predicted band size : 42kD
Exposure time : 10 second
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