Anti-Pumilio 1 antibody [EPR3795]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Immunohistochemical analysis of Pumilio 1 in paraffin embedded Human fetal kidney tissue using an unpurified ab92545 at a 1/100 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid carcinoma tissue sections labeling Pumilio 1 with Purified ab92545 at 1 : 100 dilution (1.18 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1 : 0 dilution. PBS instead of the primary antibody was used as the negative control.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Pumilio 1 with purified ab92545 at 1 : 500 dilution (0.2μg/ml). Cells were fixed in 100% Methanol. ab150077 Goat anti rabbit IgG(Alexa Fluor^® 488) was used as the secondary antibody at 1 : 1000 dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Intracellular Flow Cytometry analysis of HEK-293 (Human embryonic kidney epithelial cell) cells labeling Pumilio 1 with purified ab92545 at 1/20 dilution (10 ug/ml) (red). Cells were fixed with 4% Paraformaldehyde and permeabilized with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluorr^® 488) secondary antibody was used at 1/2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Overlay histogram showing HEK293 cells stained with an unpurified ab92545 (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 (ab92545, 1/100 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 IgG (monoclonal) (1μg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HEK293 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Immunofluorescent staining of Pumilio 1 in HeLa cells using an unpurified ab92545 at a 1/100 dilution.

• WB

Lab

Western blot - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Blocking and diluting buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-Pumilio 1 antibody [EPR3795] (ab92545) at 1/5000 dilution

Lane 1:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 126 kDa

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• WB

Lab

Western blot - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Western blot : Anti-Pumilio 1 antibody [EPR3795] ab92545 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 140 kDa in Wild-type HeLa cell lysates with no signal observed at this size in PUM1 knockout HeLa cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5% Milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Pumilio 1 antibody [EPR3795] (ab92545) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysates at 20 µg

Lane 2:

Western blot - Human PUM1 (Pumilio 1) knockout HeLa cell line (<a href='/ko/products/cell-lines/human-pum1-pumilio-1-knockout-hela-cell-line-ab265285'>ab265285</a>) at 20 µg

Lane 3:

K562 cell lysates at 20 µg

Lane 4:

HEK-293 cell lysates at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 140 kDa

Observed band size: 140 kDa

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• WB

Unknown

Western blot - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

All lanes:

Western blot - Anti-Pumilio 1 antibody [EPR3795] (ab92545) at 1/1000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

293T cell lysates at 10 µg

Secondary

All lanes:

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

Predicted band size: 126 kDa

false

• WB

Lab

Western blot - Anti-Pumilio 1 antibody [EPR3795] (AB92545)

Blocking and diluting buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-Pumilio 1 antibody [EPR3795] (ab92545) at 1/1000 dilution

Lane 1:

Mouse cerebellum tissue lysate at 20 µg

Lane 2:

Rat brain tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 126 kDa

false