Anti-IL-1 beta antibody [EPR24895-116]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunohistochemical analysis of paraffin-embedded (A) THP-1 (human monocytic leukemia monocyte) cells treated first with 80 nM TPA for overnight, then replaced with 100 ng/ml LPS for 3 hours, 300 ng/ml BFA was then added for additional 3 hours cell pellet. (B) Untreated THP-1 cell pellet. tissue labeling IL-1 beta with ab315084 at 1/8000 (0.063 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) THP-1 treated first with 80 nM TPA for overnight, then replaced with 100 ng/ml LPS for 3 hours, 300 ng/ml BFA was then added for additional 3 hours cell pellet, negative staining on (B) untreated THP-1 cell pellet.The section was incubated with ab315084 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).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunohistochemical analysis of paraffin-embedded (A) HEK-293T (human epithelial cell line from embryonic kidney) transfected with a IL1B expression vector containing a Myc-His tag. (B) HEK-293T transfected with empty vector containing a Myc-His tag. tissue labeling IL-1 beta with ab315084 at 1/8000 (0.063 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) HEK-293T (human epithelial cell line from embryonic kidney) transfected with a IL1B expression vector containing a Myc-His tag, negative staining on (B) HEK-293T transfected with empty vector containing a Myc-His tag. The section was incubated with ab315084 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).

Heat mediated antigen retrieval was performed 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 beta antibody [EPR24895-116] (AB315084)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized THP-1 (human monocytic leukemia monocyte) treated with 80nM TPA overnight and then treated with 100ng/ml LPS for 6h and 300ng/ml BFA for 3h (Red) / Untreated THP-1 (Green) cells labelling IL-1 beta with ab315084 at 1/500 dilution (0.1 ug)/Red and Green compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized THP-1 (human monocytic leukemia monocyte) cells labelling IL-1 beta with ab315084 at 1/50 (10.14 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).

Confocal image showing cytoplasmic and nuclear staining in THP-1 cells treated with TPA (80 nM) for 16 h, then LPS (100 ng/ml) for 3 h with BFA (300 ng/ml) for the last 3 h.Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).

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 (2 ug/ml) dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunohistochemical analysis of paraffin-embedded Human colon carcinoma tissue labeling IL-1 beta with ab315084 at 1/500 (1.014 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on immune cells of human colon carcinoma (PMID : 29803656). The section was incubated with ab315084 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).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunohistochemical analysis of paraffin-embedded (A) Rat lung treated with LPS (1ug/ml for 16 hours) and BFA(1ug/ml for 16 hours). (B) Untreated rat lung. tissue labeling IL-1 beta with ab315084 at 1/500 (1.014 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on immune cells of (A) rat lung treated with LPS (1ug/ml for 16 hours) and BFA(1ug/ml for 16 hours), negative staining on (B) Untreated rat lung. The section was incubated with ab315084 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).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunohistochemical analysis of paraffin-embedded (A) RAW 264.7 (mouse leukemia cells of monocyte macrophage) cells treated with 100ng/ml LPS for 4+3 hours, 1µg/ml BFA was then added for additional 3 hours cell pellet. (B) Untreated RAW 264.7 cell pellet. tissue labeling IL-1 beta with ab315084 at 1/500 (1.014 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) RAW 264.7 cells treated with 100ng/ml LPS for 4+3 hours, 1µg/ml BFA was then added for additional 3 hours cell pellet, negative staining on (B) Untreated RAW 264.7 cell pellet. The section was incubated with ab315084 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).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunohistochemical analysis of paraffin-embedded Mouse colon carcinoma tissue labeling IL-1 beta with ab315084 at 1/500 (1.014 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on immune cells of mouse colon carcinoma (PMID : 26292622). The section was incubated with ab315084 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).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Immunohistochemical analysis of paraffin-embedded (A) NR8383 (rat alveolar macrophages) cells treated with 100ng/ml LPS for 4+3 hours, 1 ug/ml BFA was then added for additional 3 hours cell pellet. (B) Untreated NR8383 cell pellet. tissue labeling IL-1 beta with ab315084 at 1/2000 (0.254 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) NR8383 cells treated with 100ng/ml LPS for 4+3 hours, 1 ug/ml BFA was then added for additional 3 hours cell pellet, negative staining on (B) Untreated NR8383 cell pellet. The section was incubated with ab315084 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).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• WB

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Western blot - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Blocking and diluting buffer and concentration : Intercept® (TBS) Blocking Buffer diluted with an equal volume of TBS.

The samples were run on a Bis-Tris gel under reducing conditions.

Western blot : Anti-IL-1 beta antibody (ab315084) staining at 1/1000 dilution, shown in green; Mouse anti-alpha Tubulin antibody [DM1A] (ab7291) loading control staining at 1/20, 000 dilution, shown in red.

In Western blot, ab315084 was shown to bind specifically to IL-1 beta. Target of interest was observed at 34, 17 kDa in Wild-type THP-1 treated first with 80 nM TPA for overnight, then replaced with 100 ng/ml LPS for 3 hours, 300 ng/ml BFA was then added for additional 3 hours, whole cell lysate (lane 2) with no signal observed at this size in IL-1 beta knockout THP-1 treated first with 80 nM TPA for overnight, then replaced with 100 ng/ml LPS for 3 hours, 300 ng/ml BFA was then added for additional 3 hours cell line ab273762 (knockout cell lysate ab275508) (lane 4). To generate this image, samples were first run on an SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in a fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4 °C. Blots were washed in TBS-T, incubated with secondary antibodies Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution for 1 h at room temperature, washed again then imaged.

All lanes:

Western blot - Anti-IL-1 beta antibody [EPR24895-116] (ab315084) at 1/1000 dilution

Lane 1:

Untreated Wild-type THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Lane 2:

Wild-type THP-1 treated first with 80 nM TPA for overnight, then replaced with /ml LPS for 3 hours, 300 ng/ml BFA was then added for additional 3 hours, whole cell lysate at 20 µg

Lane 3:

Untreated IL-1 beta knockout THP-1 whole cell lysate at 20 µg

Lane 4:

IL-1 beta knockout THP-1 treated first with 80 nM TPA for overnight, then replaced with /ml LPS for 3 hours, 300 ng/ml BFA was then added for additional 3 hours, whole cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG H&L (800CW) and Goat Anti-Mouse IgG H&L (680RD) at 1/20000 dilution

Observed band size: 34 kDa,17 kDa

false

• WB

Supplier Data

Western blot - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

Exposure time : Lanes 1-2 : 1 second; Lanes 3-4 : 6 seconds.

All lanes:

Western blot - Anti-IL-1 beta antibody [EPR24895-116] (ab315084) at 1/1000 dilution

Lane 1:

Untreated THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Lane 2:

THP-1 treated first with 80 nM TPA for overnight, then replaced with /ml LPS for 3 hours, 300 ng/ml BFA was then added for additional 3 hours, whole cell lysate at 20 µg

Lane 3:

Untreated RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 4:

RAW 264.7 treated with /ml LPS for 4 hours, 1/ml BFA was then added for additional 3 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 1/100000 dilution

Observed band size: 34 kDa

false

• WB

Supplier Data

Western blot - Anti-IL-1 beta antibody [EPR24895-116] (AB315084)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-IL-1 beta antibody [EPR24895-116] (ab315084) at 1/1000 dilution

Lane 1:

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

Lane 2:

NR8383 treated with 100 ng/ml LPS for 4 hours, 1 μg/ml BFA was then added for additional 3 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: 34 kDa

false

Exposure time: 6s