Anti-STING antibody [EPR13130-55]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-STING antibody [EPR13130-55] (AB239074)

Immunohistochemical analysis of formalin fixed paraffin (FFPE) embedded tonsil labelling STING with ab239074 at a concentration of 0.01µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with an OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was conducted for 32 min with DISCOVERY cell conditioning solution (CC1)  100°C, pH 8.5. ab239074 was incubated at 37°C for 16min. Sections were counterstained is with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-STING antibody [EPR13130-55] (AB239074)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized THP-1 (human monocytic leukemia cell line) cell line labeling TMEM173 with ab239074 at 1/500 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-STING antibody [EPR13130-55] (AB239074)

Immunohistochemical analysis of paraffin-embedded human colon cancer tissue labeling TMEM173 with ab239074 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Positive staining in human colon cancer (PMID : 26748708) is observed. Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-STING antibody [EPR13130-55] (AB239074)

Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling TMEM173 with ab239074 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Positive staining in human tonsil (PMID : 28874664) is observed. Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-STING antibody [EPR13130-55] (AB239074)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized THP-1 (human monocytic leukemia cell line) cells labeling TMEM173 with ab239074 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic and membranous staining in THP-1 cell line is observed.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-STING antibody [EPR13130-55] (AB239074)

Immunohistochemical analysis of paraffin-embedded human prostate hyperplasia tissue labeling TMEM173 with ab239074 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Positive staining in basal cells of human prostate hyperplasia is observed. Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-STING antibody [EPR13130-55] (AB239074)

Tissue Microarrays stained for "Anti-STING antibody [EPR13130-55]" using "ab239074"in immunohistochemical analysis. This table provides a detailed overview of positive (tick mark) and negative (cross mark) staining per sample type tested. The sections were pre-treated using Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). The sections were incubated with ab239074 at +4°C overnight followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer).

• IP

Unknown

Immunoprecipitation - Anti-STING antibody [EPR13130-55] (AB239074)

TMEM173 was immunoprecipitated from 0.35 mg of THP-1 (human monocytic leukemia cell line) whole cell lysate with ab239074 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab239074 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

Lane 1 : THP-1 whole cell lysate 10 μg (Input).

Lane 2 : ab239074 IP in THP-1 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab239074 in THP-1 whole cell lysate.

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

Exposure time : 30 seconds.

All lanes:

Immunoprecipitation - Anti-STING antibody [EPR13130-55] (ab239074)

Predicted band size: 42 kDa

false

• IP

Lab

Immunoprecipitation - Anti-STING antibody [EPR13130-55] (AB239074)

Immunoprecipitation of STING1 in THP-1 cells. Lysates were prepared and immunoprecipitation was performed using 2 µg of ab239074 pre-coupled to Protein A beads. Samples were then washed and processed for western blot.

These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Immunoprecipitation - Anti-STING antibody [EPR13130-55] (ab239074) at 2 µg

All lanes:

THP-1 cells

false

• WB

Lab

Western blot - Anti-STING antibody [EPR13130-55] (AB239074)

Lanes 1 - 4 : Merged signal (red and green). Green - ab239074 observed at 37 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab239074 was shown to react with TMEM173 in wild-type THP-1 cells in Western blot with loss of signal observed in TMEM173 knockout cell line ab270493 (TMEM173 knockout cell lysae ab270516). Wild-type THP-1 and TMEM173 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab239074 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-STING antibody [EPR13130-55] (ab239074) at 1/1000 dilution

Lane 1:

Wild-type THP-1 cell lysate at 20 µg

Lane 2:

TMEM173 knockout THP-1 cell lysate at 20 µg

Lane 2:

Western blot - Human TMEM173 knockout THP-1 cell line (<a href='/en-us/products/cell-lines/human-tmem173-knockout-thp-1-cell-line-ab270493'>ab270493</a>)

Lane 3:

Human Tonsil tissue lysate at 20 µg

Lane 4:

Human Thymus tissue lysate at 20 µg

Predicted band size: 42 kDa

Observed band size: 37 kDa

false

• WB

Supplier Data

Western blot - Anti-STING antibody [EPR13130-55] (AB239074)

Blocking and diluting buffer and concentration : 1% BSA/TBST.

Performed under reducing conditions.

In Western blot, ab324229 was shown to bind specifically to STING (phospho S366). Target of interest was observed at 40kDa in wild-type THP-1 cell lysates (lane 2) with no signal observed at this size in STING knockout cell line (lane 3-4) (lane 3, knockout cell line ab270493 / knockout cell lysate ab270516).

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

In Western blot, Anti-STING antibody [EPR13130-55] (ab239074) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-STING (phospho S366) antibody [EPR29040-93] (<a href='/en-us/products/primary-antibodies/sting-phospho-s366-antibody-epr29040-93-ab324229'>ab324229</a>) at 1/1000 dilution

Lane 1:

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

Lane 2:

Wild-type THP-1 treated first with 80nM TPA for 24h, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, whole cell lysate (untreated membrane) at 20 µg

Lane 3:

Untreated STING knockout THP-1 whole cell lysate (untreated membrane) at 20 µg

Lane 4:

STING knockout THP-1 treated first with 80nM TPA for 24h, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, whole cell lysate (untreated membrane) at 20 µg

Lane 5:

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

Lane 6:

Wild-type THP-1 treated first with 80nM TPA for 24h, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, whole cell lysate (alkaline phosphatase treated membrane) at 20 µg

Lane 7:

Untreated STING knockout THP-1 whole cell lysate (alkaline phosphatase treated membrane) at 20 µg

Lane 8:

STING knockout THP-1 treated first with 80nM TPA for 24h, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, whole cell lysate (alkaline phosphatase treated membrane) 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: 40 kDa,124 kDa

false

Exposure time: 59s

• WB

Supplier Data

Western blot - Anti-STING antibody [EPR13130-55] (AB239074)

Blocking and diluting buffer and concentration : 1% BSA/TBST.

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

In Western blot, Anti-STING antibody [EPR13130-55] (ab239074) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-STING (phospho S366) antibody [EPR29040-93] (<a href='/en-us/products/primary-antibodies/sting-phospho-s366-antibody-epr29040-93-ab324229'>ab324229</a>) 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 80nM TPA for 24 hours, then change fresh medium, transfect 10ug/ml poly(dA:dT) for 6 hours, whole cell lysate at 20 µg

Lane 3:

THP-1 treated first with 80nM TPA for 24 hours, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, whole cell lysate at 20 µg

Lane 4:

THP-1 treated first with 80nM TPA for 24 hours, then change fresh medium, transfect 10ug/ml poly(dA:dT) for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, whole cell lysate at 20 µg

Lane 5:

THP-1 treated first with 80nM TPA for 24 hours, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, 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: 40 kDa,124 kDa

false

Exposure time: 59s

• WB

Unknown

Western blot - Anti-STING antibody [EPR13130-55] (AB239074)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-STING antibody [EPR13130-55] (ab239074) at 1/1000 dilution

Lane 1:

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

Lane 2:

HT1080 (human fibrosarcoma cell line) whole cell lysate at 20 µg

Lane 3:

Human tonsil 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

Predicted band size: 42 kDa

Observed band size: 36 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-STING antibody [EPR13130-55] (AB239074)

ab239074 was shown to react with STING1 in wild-type THP-1 cells in Western blot with loss of signal observed in STING1 knockout cell line ab270493. Wild-type THP-1 and STING1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab239074 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-STING antibody [EPR13130-55] (ab239074) at 1/1000 dilution

Lane 1:

Wild-type THP-1 lysate at 40 µg

Lane 2:

STING1 knock-out THP-1 lysate at 40 µg

false

• WB

Supplier Data

Western blot - Anti-STING antibody [EPR13130-55] (AB239074)

Blocking and diluting buffer and concentration : 1% BSA/TBST.

Anti-Vinculin antibody [EPR8185] (ab129002) (1/10000) (124KDa); Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) (1/5000);

In Western blot, Anti-STING antibody [EPR13130-55] (ab239074) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-STING (phospho S366) antibody [EPR29040-93] (<a href='/en-us/products/primary-antibodies/sting-phospho-s366-antibody-epr29040-93-ab324229'>ab324229</a>) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) cells transfected with an empty vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human wild-type STING expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 3:

293T cells transfected with a human wild-type STING expression vector containing a myc-His-tag® treated first with 80nM TPA for 24h, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, whole cell lysate at 20 µg

Lane 4:

293T cells transfected with a human STING (S366A mutation) expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 5:

293T cells transfected with a human STING (S366A mutation) expression vector containing a myc-His-tag® treated first with 80nM TPA for 24h, then change fresh medium, transfect 10ug/ml poly(dA:dT) and treated with 10 uM MG-132 for 6 hours, 100 uM Calyculin A was then added for additional 30 minutes, 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: 40 kDa,124 kDa

false

Exposure time: 59s