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Tissue/ cell preparation (Human). Fibroblast cells (HT1080).
This monoclonal antibody to integrin beta 1 has been knockout validated in ICC/IF and flow cytometry. The expected signal was observed in wild type cells and no signal was seen in knockout cells.
Alternative versions available:
This antibody inhibits the function of all beta 1 Integrins.
Our Abpromise guarantee covers the use of ab24693 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Blocking||Use at an assay dependent concentration.|
|ICC/IF||Use a concentration of 1 - 5 µg/ml.|
|Flow Cyt||Use 0.1-1µg for 106 cells.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|WB||1/1000. Predicted molecular weight: 88 kDa.|
ab24693 staining Integrin beta 1 in wild-type HAP1 cells (top panel) and Integrin beta 1 knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10min), 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 with ab24693 at 5μg/ml concentration and ab202272 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Mouse IgG (Alexa Fluor® 488) (ab150117) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.
Overlay histogram showing HAP1 wildtype (green line) and HAP1-ITGB1 knockout cells (red line) stained with ab24693. Live HAP1 wildtype and HAP1-ITGB1 knockout cells were incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab24693, 0.1µg/0.5x106 cells) for 30 min at 22°C. A mouse IgG1 isotype control antibody (ab170190) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-ITGB1 knockout - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity). Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.
Overlay histogram showing MCF7 cells stained with ab24693 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab24693, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was a goat anti-mouse DyLight® 488 (IgG; H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in MCF7 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
ICC/IF image of ab24693 stained HeLa cells. The cells were formaldehyde fixed, permeabilization with 0.5% TritonX/PBS and then blockedwith 5% BSA for 30 minutes at 21°C. The cells were then incubated with the antibody (ab24693, 1/200) for 1 hour at 21°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/500 dilution. DAPI was used to stain the cell nuclei (blue) and Phalloidin-TRITC (red) the actin cytoskeleton.