Anti-STING antibody [EPR13130] - BSA and Azide free

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

Intracellular Flow Cytometry analysis of THP1 cells using unpurified ab181125 at a 1/10 dilution (red) or a Rabbit monoclonal IgG (negative) (green). Goat anti rabbit IgG (FITC) secondary used at a 1/150 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab181125).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

Clone EPR13130 (ab227128) has been successfully conjugated by Abcam. This image was generated using Anti-TMEM173 antibody [EPR13130] (PE). Please refer to ab208874 for protocol details.

Overlay histogram showing HeLa cells stained with ab208874 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab208874 1/500 dilution) for 30 min at 22°C.Isotype control antibody (black line) was rabbit IgG (monoclonal) Phycoerythrin (ab209478) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control. Acquisition of >5000 events were collected using a 50 mW Yellow/Green laser (561nm) and 586/15 bandpass filter. This antibody gave a positive signal in HeLa cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Triton X-100 for 15 min used under the same conditions.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

Clone EPR13130 (ab227128) has been successfully conjugated by Abcam. This image was generated using Anti-TMEM173 antibody [EPR13130] (Alexa Fluor® 488). Please refer to ab198950 for protocol details.

ab198950 staining TMEM173 in HeLa cells. The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab198950 at 1/200 dilution (shown in green) and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 594), at 1/250 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

Overlay histogram showing THP-1 cells fixed in 4% PFA and stained with purified ab181125 at a dilution of 1 in 20 (red line). The secondary antibody used was FITC goat anti-rabbit at a dilution of 1 in 500. Rabbit monoclonal IgG was used as an isotype control (black line) and cells incubated in the absence of both primary and secondary antibody were used as a negative control (blue line). This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab181125).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

Immunofluorescence staining of HeLa cells with purified ab181125 at a working dilution of 1/1000, counter-stained with DAPI. The secondary antibody was Alexa Fluor^® 488 goat anti-rabbit (ab150077), used at a dilution of 1/1000. ab7291, a mouse anti-tubulin antibody (1/1000), was used to stain tubulin along with ab150120 (Alexa Fluor^® 594 goat anti-mouse, 1/1000), shown in the top right hand panel. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative controls are shown in bottom middle and right hand panels - for negative control 1, purified ab181125 was used at a dilution of 1/500 followed by an Alexa Fluor^® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500. For negative control 2, ab7291 (mouse anti-tubulin) was used at a dilution of 1/500 followed by an Alexa Fluor^® 488 goat anti-rabbit antibody (ab150077) at a dilution of 1/400.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab181125).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

Clone EPR13130 (ab227128) has been successfully conjugated by Abcam. This image was generated using Anti-TMEM173 antibody [EPR13130] (Alexa Fluor® 647). Please refer to ab198952 for protocol details.

ab198952 staining TMEM173 in HeLa cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab198952 at 1/100 dilution (shown in red) and ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 488), at 1/250 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

This product also gave a positive signal under the same testing conditions in HeLacells fixed with 4% formaldehyde (10 min).

• IP

Lab

Immunoprecipitation - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

This data was developed using ab181125, the same antibody clone in a different buffer formulation.

Immunoprecipitation of STING1 in THP-1 cells. Lysates were prepared and immunoprecipitation was performed using 2 µg of ab181125 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] (<a href='/en-us/products/primary-antibodies/sting-antibody-epr13130-ab181125'>ab181125</a>) at 2 µg

All lanes:

THP-1 cells

false

• WB

Lab

Western blot - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

This data was developed using ab181125, the same antibody clone in a different buffer formulation.

ab181125 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 ab181125 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] (<a href='/en-us/products/primary-antibodies/sting-antibody-epr13130-ab181125'>ab181125</a>) 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

Lab

Western blot - Anti-STING antibody [EPR13130] - BSA and Azide free (AB227128)

False colour image of Western blot : Anti-STING antibody [EPR13130] staining at 1/1000 dilution, shown in green; loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) at 1/20000 dilution, shown in red. In Western blot, ab181125 was shown to bind specifically to STING. A band was observed at 37 kDa in wild-type THP-1 cell lysates with no signal observed at this size in TMEM173 knockout cell line ab270493 (knockout cell lysate ab270516). To generate this image, wild-type and TMEM173 knockout THP-1 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution 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 IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab181125).

All lanes:

Western blot - Anti-STING antibody [EPR13130] (<a href='/en-us/products/primary-antibodies/sting-antibody-epr13130-ab181125'>ab181125</a>) 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 cell lysate at 20 µg

Lane 4:

Human Thymus cell lysate at 20 µg

Predicted band size: 42 kDa

Observed band size: 37 kDa

false