Recombinant 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.

Blocking/Dilution buffer: 5% NFDM/TBST.

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.

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.

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.

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.

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.

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. 

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).