Western blot - Integrin beta 1 antibody [EP1041Y] (ab52971)

Anti-Integrin beta 1 antibody [EP1041Y] - Carboxyterminal end (ab52971) at 1/500 dilution + U937 cell lysate at 10 µg

Secondary
goat anti-rabbit HRP labelled at 1/2000 dilution

Predicted band size : 88 kDa
Observed band size : 140 kDa (why is the actual band size different from the predicted?)

Immunohistochemistry (Paraffin-embedded sections) - Integrin beta 1 antibody [EP1041Y] (ab52971)

ab52971, at a 1/250 dilution, staining Human Integrin beta 1 in breast carcinoma using Immunohistochemistry, Paraffin Embedded tissue.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Integrin beta 1 antibody [EP1041Y] - Carboxyterminal end (ab52971)

ab52971 staining human breast cancer metastasis tissue sections by IHC-P.  Sections were formaldehyde fixed and subjected to heat mediated antigen retrieval in citrate buffer (pH 6) prior to blocking with a commercial blocking reagent and incubation with the antibody (diluted 1/100) for 18 hours at 4°C.  A HRP-conjugated goat anti-rabbit was used as the secondary antibody.  This image shows a cancer metastasis at 40x with beta1 staining (in red) in both blood vessels and tumour cells. Blue is Hoechst for nuclei.  The antibody detection was enhanced using a commercial Cy3 tyramide signal amplification kit.

This image is courtesy of an anonymous Abreview

Immunocytochemistry/ Immunofluorescence - Integrin beta 1 antibody [EP1041Y] - Carboxyterminal end (ab52971)

ICC/IF image of ab52971 stained Hek293 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab52971, 1/1000 dilution) overnight at +4ºC. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

Flow Cytometry - Integrin beta 1 antibody [EP1041Y] - Carboxyterminal end (ab52971)

Overlay histogram showing MCF-7 cells stained with ab52971 (red line). The cells were fixed with methanol (5 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 (ab52971, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit monoclonal IgG (1µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in MCF-7 cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

Immunocytochemistry/ Immunofluorescence - Anti-Integrin beta 1 antibody [EP1041Y] - Carboxyterminal end (ab52971)

ab52971 at a 1/400 dilution staining Integrin beta 1 in rat medullary thyroid carcinoma cells cells by Immunocytochemistry/ Immunofluorescence. The cells were fixed with 4% paraformaldehyde for 15 minutes, blocked for 30 minutes using 5% goat serum (nonpermeabilized). The secondary antibody used was a DyLight 549-conjugated goat anti-rabbit at a 1/500 dilution.

Image from Tharmalingam S et al, J Biol Chem. 2011 Nov 25;286(47):40922-33. Epub 2011 Oct 3, Fig 2. DOI 10.1074/jbc.M111.265454