重组Anti-COX2 / Cyclooxygenase 2抗体[EPR12012] (ab179800)

Compared with ab179800, ab283574 has higher sensitivity, we recommend ab283574 as an alternative for testing COX2 in western blot.

ab181602 was used as a loading control at a 1/1000000 dilution.

Blocking and dilution buffer: 5% NFDM/TBST.

Exposure time: lane 1: 180 seconds, lane 2: 140 seconds. We recommend using a higher sensitive ECL substrate to increase the band intensity

Western blot: Anti-PTGS2 antibody [EPR12012] (ab179800) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab179800 was shown to bind specifically to PTGS2. A band was observed at 69 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in PTGS2 knockout cell line. To generate this image, wild-type and PTGS2 knockout RAW 264.7 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween® 20 (TBS-T) 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

Blocking buffer and concentration: 5% NFDM/TBST 

Diluting buffer and concentration: 5% NFDM/TBST

COX2 is expressed at a low level in Raw264.7, mouse retina, hippocampus, heart, kidney etc. (PMID: 22015457, PMID: 26001832, PMID: 23045674, PMID: 33737575).

The WB is using a higher sensitivity ECL substrate.

False colour image of Western blot: Anti-COX2 / Cyclooxygenase 2 antibody [EPR12012] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab179800 was shown to bind specifically to COX2 / Cyclooxygenase 2. A band was observed at 75 kDa in wild-type A549 cell lysates with no signal observed at this size in PTGS2 knockout cell line ab280802 (knockout cell lysate ab283825). To generate this image, wild-type and PTGS2 knockout A549 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.

Immunocytochemistry/ Immunofluorescence analysis of U-87 MG (human glioblastoma-astrocytoma epithelial cell) cells labeling COX2 / Cyclooxygenase 2 with ab179800 at 1/50 dilution. ab150077 (AlexaFluor^®488 Goat anti-Rabbit) at 1/1000 was used as secondary antibody. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100. ab195889, Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1/200 was used as counterstain. Nuclie were stained blue with DAPI.

Confocal image showing cytoplasmic staining in U-87 MG cell line.
Negative control: MCF7 （PMID: 18199541）

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colonic carcinoma tissue labelling COX2 / Cyclooxygenase 2 with unpurified ab179800 at a dilution of 1/250.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded) analysis of human colon tissue labeling COX2 / Cyclooxygenase 2 with unpurified ab179800 at a dilution of 1/250.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded) analysis of human liver tissue labelling COX2 / Cyclooxygenase 2 with unpurified ab179800 at a dilution of 1/250.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat kidney tissue labelling COX2 / Cyclooxygenase 2 with purified ab179800 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human lung carcinoma tissue labelling COX2 / Cyclooxygenase 2 with purified ab179800 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

ab179800 (purified) at 1/30 immunoprecipitating COX2 in A549 whole cell lysate.

Lane 1 (input): A549 whole cell lysate (10µg)

Lane 2 (+): ab179800 + A549 whole cell lysate.

Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab179800 in A549 whole cell lysate.

For western blotting, HRP-conjugated anti-rabbit IgG, specific for the reduced form of IgG, was used as the secondary antibody (1/1500).

Blocking buffer and concentration: 5% NFDM/TBST.

Diluting buffer and concentration: 5% NFDM /TBST.

Western blot analysis on immunoprecipitation pellet from A549 cell lysate using unpurified ab179800.