Anti-beta Arrestin 1 antibody [E274] (ab32099) at 1/1000 dilution (purified) + Jurkat whole cell lysate at 20 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 50 kDa
Observed band size: 50 kDa



Blocking and dilution buffer: 5% NFDM/TBST

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human lymph node tissue labelling beta Arrestin 1 with unpurified ab32099 at a dilution of 1/100.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue labelling beta Arrestin 1 with purified ab32099 at a dilution of 1/250. Heat mediated antigen retrieval was performed using EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

Immunocytochemistry/Immunofluorescence analysis of PC-3 cells labelling beta Arrestin 1 with purified ab32099 at a dilution of 1/150. 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 (1/150) and secondary antibody, ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/1000).

Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000).

Overlay histogram showing PC3 cells stained with unpurified ab32099 (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 (unpurified ab32099, 1/10 dilution) for 30 min at 22°C. The secondary antibody used was DyLight^® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. 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 decreased signal in PC3 cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

All lanes : Anti-beta Arrestin 1 antibody [E274] (ab32099) at 1/2000 dilution (purified)

Lane 1 : HEK293 whole cell lysate
Lane 2 : SH-SY5Y whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 50 kDa
Observed band size: 50 kDa



Blocking and dilution buffer: 5% NFDM/TBST

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse cerebral cortex tissue labelling beta Arrestin 1 with purified ab32099 at a dilution of 1/250. Heat mediated antigen retrieval was performed using EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

All lanes : Anti-beta Arrestin 1 antibody [E274] (ab32099) at 1/1000 dilution (purified)

Lane 1 : C2C12 whole cell lysate
Lane 2 : PC-12 whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 50 kDa
Observed band size: 50 kDa



Blocking and dilution buffer: 5% NFDM/TBST

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat cerebral cortex tissue labelling beta Arrestin 1 with purified ab32099 at a dilution of 1/250. Heat mediated antigen retrieval was performed using EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

Unpurified ab32099 staining beta Arrestin 1 in C4-2B (Human prostate cancer cell line) by ICC/IF (Immunocytochemistry/immunofluorescence).
Cells were fixed with paraformaldehyde and blocked with 1% serum for 1 hour at 21°C. Samples were incubated with primary antibody (1/100 in diluent) for 1 hour at 21°C. An Alexa Fluor^® 488-conjugated goat anti-rabbit polyclonal IgG (1/200) was used as the secondary antibody.

See Abreview

Unpurified ab32099 staining beta Arrestin 1 in human prostate carcinoma tissue section by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue underwent paraformaldehyde fixation before heat mediated antigen retrieval in Tris/EDTA pH9.0 and then blocking with 1% donkey serum for 1 hour at 20°C was performed. The primary antibody was diluted 1/100 and incubated with sample for 1 hour at 20°C in PBS. A Biotin conjugated donkey polyclonal to rabbit IgG was used undiluted as secondary antibody.

See Abreview