Anti-IL-1 alpha antibody [EPR25263-27]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-IL-1 alpha antibody [EPR25263-27] (AB300499)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized THP-1 (human monocytic leukemia monocyte) cells labelling IL-1 alpha with ab300499 at 1/50 (10.5 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2ug/mL dilution (Green). Confocal image showing increased nuclear staining in THP-1 cells treated with Phorbol-12-myristate-13-acetate (50 nM) for 24 h. is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2ug/mL dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 alpha antibody [EPR25263-27] (AB300499)

Immunohistochemical analysis of paraffin-embedded A Mouse lung treate tissue labeling IL-1 alpha with ab300499 at 1/500 (1.05 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Positive staining on mouse lung induced by LPS (A) and no staining on untreated mouse lung. The section was incubated with ab300499 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin. Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-IL-1 alpha antibody [EPR25263-27] (AB300499)

Flow Cytometry (Intracellular) analysis of RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) treated with 10 µg/ml LPS for 24 hours (Right) / Untreated control (Left) IL-1 alpha with ab300499 at a 1/500 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A Goat Anti-Rabbit IgG (Alexa Fluor® 488, (ab150081)) was used as the secondary antibody at a 1/2000 dilution. The expression level of IL-1 alpha was upregulated by LPS stimulation (PMID : 25870118).

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-IL-1 alpha antibody [EPR25263-27] (AB300499)

Flow Cytometry (Intracellular) analysis of mouse bone marrow cells treated with 5 µg/ml LPS for 7 hours (Right) / Untreated control (Left) labelling IL-1 alpha with ab300499 at a 1/500 dilution. Cells were fixed with 2% paraformaldehyde and permeabilized using True-Nuclear buffer. A Goat Anti-Rabbit IgG (Alexa Fluor® 488, (ab150081)) was used as the secondary antibody at a 1/2000 dilution. Cells were surface stained with BV421 conjugated CD11b, then fixed with PFA followed by intracellularly stained with ab300499.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-IL-1 alpha antibody [EPR25263-27] (AB300499)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) cells labelling IL-1 alpha with ab300499 at 1/50 (10.5 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2ug/mL dilution (Green). Confocal image showing increased nuclear staining in Raw 264.7 cells treated with lipopolysaccharide (10 ug/ml) for 24 h (PMID : 25870118) is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2ug/mL dilution.

• WB

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Western blot - Anti-IL-1 alpha antibody [EPR25263-27] (AB300499)

Blocking and diluting buffer and concentration : 5% NFDM/TBST The expression level of IL-1 alpha was upregulated by LPS stimulation (PMID : 25870118)

All lanes:

Western blot - Anti-IL-1 alpha antibody [EPR25263-27] (ab300499) at 1/1000 dilution

Lane 1:

Untreated RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate

Lane 2:

RAW264.7 treated with /ml lipopolysaccharide (LPS) for 7 hours, whole cell lysate 40 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 30 kDa

false

Exposure time: 59s