Anti-IL-1 beta antibody [EPR16805-15]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-IL-1 beta antibody [EPR16805-15] (AB234437)

Immunohistochemical analysis of 4% paraformaldehyde fixed, 0.1% TritonX-100 permeabilised J774A.1 (mouse reticulum cell sarcoma monocyte macrophage) cells labeling IL-1 beta with ab234437 at 1/50 (10μg/ml) dilution. ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was used as a secondary antibody at 1/1000 (2μg/ml) dilution. Counterstained with ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594). Nuclear staining : DAPI.

Confocal image showing cytoplasmic staining in J774A.1 cells treated with Lipopolysaccharides (100 ng/ml) for 4h, with addition of brefeldin A (1 μg/ml) for the last 3 hours.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-IL-1 beta antibody [EPR16805-15] (AB234437)

Intracellular flow cytometric analysis of4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) cells that were either treated (100 ng/ml LPS for 3h, then add 300 ng/ml Brefeldin A for 3h labeling) (red) or untreated (green) labeling IL-1 beta with ab234437 at 1/60 dilution compared with a rabbit monoclonal IgG Isotype control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti-Rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

IL-1 beta is not present under homeostatic conditions; it is induced and secreted only upon inflammatory signals and its secretion is tightly controlled at the levels of transcription, mRNA stability, translation, post-translational modifications and processing (PMID : 26686225).

• IP

Unknown

Immunoprecipitation - Anti-IL-1 beta antibody [EPR16805-15] (AB234437)

IL-1 beta was immunoprecipitated from 0.35 mg of RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) (treated with 100 ng/ml LPS for 3h, then add 300 ng/ml Brefeldin A for 3h) whole cell lysate with ab234437 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab234437 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used as secondary antibody at 1/5000 dilution.

Lane 1 : RAW 264.7 (treated with 100 ng/ml LPS for 3h, then add 300 ng/ml Brefeldin A for 3h) whole cell lysate 10 μg (Input).
Lane 2 : ab234437 IP in RAW 264.7 (treated with 100 ng/ml LPS for 3h, then add 300 ng/ml Brefeldin A for 3h) whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab234437 in RAW 264.7 (treated with 100 ng/ml LPS for 3h, then add 300 ng/ml Brefeldin A for 3h) whole cell lysate.

IL-1 beta is not present under homeostatic conditions; it is induced and secreted only upon inflammatory signals and its secretion is tightly controlled at the levels of transcription, mRNA stability, translation, post-translational modifications and processing (PMID : 26686225).

Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-IL-1 beta antibody [EPR16805-15] (ab234437)

Predicted band size: 30 kDa

Observed band size: 31 kDa

false

• WB

Unknown

Western blot - Anti-IL-1 beta antibody [EPR16805-15] (AB234437)

Blocking and dilution buffer : 5% NFDM/TBST.

The molecular weight observed is consistent with what has been described in the literature (PMID : 8446594).

IL-1 beta is not present under homeostatic conditions; it is induced and secreted only upon inflammatory signals and its secretion is tightly controlled at the levels of transcription, mRNA stability, translation, post-translational modifications and processing (PMID : 26686225).

All lanes:

Western blot - Anti-IL-1 beta antibody [EPR16805-15] (ab234437) at 1/1000 dilution

Lane 1:

Untreated RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 10 µg

Lane 2:

RAW 264.7 (treated with 100 ng/ml lipopolysaccharide (LPS) for 6 hours, then with 300 ng/ml Brefeldin A (BFA) added after 3 hours) whole cell lysate at 10 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

Predicted band size: 30 kDa

Observed band size: 17 kDa,28 kDa,31 kDa

false

Exposure time: 15s

• WB

Lab

Western blot - Anti-IL-1 beta antibody [EPR16805-15] (AB234437)

Blocking and dilution buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-IL-1 beta antibody [EPR16805-15] (ab234437) at 1/1000 dilution

Lanes 1 and 3:

Untreated NR8383 (rat lung macrophage (alveolar)) whole cell lysate at 20 µg

Lane 2:

NR8383 (rat lung macrophage (alveolar)) treated with 100ng/ml IFN gamma and 10ng TNF alpha for 32 hours, then with 5 µg/ml Brefeldin A (BFA) added for 6 hours whole cell lysate at 20 µg

Lane 4:

NR8383 (rat lung macrophage (alveolar)) treated with 100ng/ml LPS and 300ng /ml Brefeldin A (BFA) for 6 hours whole cell lysate at 20 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 1/20000 dilution

Observed band size: 20 kDa,31 kDa

false

Exposure time: 40s