Anti-COX1 / Cyclooxygenase 1 抗体 [EPR5866]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Unpurified ab109025 at 1/100 dilution staining COX1 / Cyclooxygenase 1 in HeLa cells by Immunofluorescence.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Unpurified ab109025 at 1/250 dilution staining COX1 / Cyclooxygenase 1 in human skin by immunohistochemistry, paraffin-embedded tissue.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cerebrum tissue sections labeling COX1 / Cyclooxygenase 1 with Purified ab109025 at 1 : 150 dilution. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1 : 0 dilution.

PBS instead of the primary antibody was used as the negative control.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Intracellular Flow Cytometry analysis of NIH/3T3 (Mouse embryonic fibroblast) cells labeling COX1 / Cyclooxygenase 1 with purified ab109025 at 1/100 dilution (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluorr^® 488) secondary antibody was used at 1/2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Immunocytochemistry/Immunofluorescence analysis of Neuro-2a (mouse neuroblastoma) labelling COX1 with purified ab109025 at 1/50. Cells were fixed with 100% methanol. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody (ab150077). Nuclei counterstained with DAPI (blue).

Control : PBS only.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat kidney tissue sections labeling COX1 / Cyclooxygenase 1 with Purified ab109025 at 1 : 150 dilution. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1 : 0 dilution.

PBS instead of the primary antibody was used as the negative control.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Overlay histogram showing NIH/3T3 cells stained with unpurified ab109025 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab109025 1/100 dilution) for 30 min at 22°C. The secondary antibody used was DyLight^® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1μg/1x10⁶ cells) used under the same conditions. Acquisition of >5000 events was performed.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse kidney tissue sections labeling COX1 / Cyclooxygenase 1 with Purified ab109025 at 1 : 150 dilution. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1 : 0 dilution.

PBS instead of the primary antibody was used as the negative control.

• IP

Lab

Immunoprecipitation - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

ab109025 (purified) at 1 : 20 dilution (0.8μg) immunoprecipitating COX1 / Cyclooxygenase 1 in C2C12 whole cell lysate.

Lane 1 (input) : C2C12 (Mouse myoblasts myoblast) whole cell lysate,10μg
Lane 2 (+) : ab109025 & C2C12 whole cell lysate
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab109025 in C2C12 whole cell lysate

For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1 : 1000 dilution.

Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (ab109025)

Predicted band size: 68 kDa

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• WB

Lab

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

False colour image of Western blot : Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] 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, ab109025 was shown to bind specifically to COX1 / Cyclooxygenase 1. A band was observed at 70 kDa in wild-type A431 cell lysates with no signal observed at this size in PTGS1 knockout cell line ab270477 (knockout cell lysate ab270500). To generate this image, wild-type and PTGS1 knockout A431 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.

Lanes 1 - 4:

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] - BSA and Azide free (<a href='/products/primary-antibodies/cox1-cyclooxygenase-1-antibody-epr5866-bsa-and-azide-free-ab219375'>ab219375</a>) at 1/1000 dilution

Lanes 1 - 4:

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (ab109025) at 1/1000 dilution

Lane 1:

Wild-type A431 cell lysate at 20 µg

Lane 2:

Western blot - Human PTGS1 knockout A-431 cell lysate (ab270500)

Lane 2:

PTGS1 knockout A431 cell lysate at 20 µg

Lane 2:

Western blot - Human PTGS1 knockout A-431 cell line (<a href='/products/cell-lines/human-ptgs1-knockout-a-431-cell-line-ab270477'>ab270477</a>)

Lane 3:

C2C12 cell lysate at 20 µg

Lane 4:

MOLT-4 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: 70 kDa

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• WB

Lab

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (ab109025) at 1/2000 dilution

Lane 1:

HaCaT (Human skin keratinocyte) whole cell lysates at 20 µg

Lane 2:

NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 68 kDa

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• WB

Lab

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (ab109025) at 1/10000 dilution

Lane 1:

A431 (Human epidermoid carcinoma epithelial cell) whole cell lysates at 20 µg

Lane 2:

L6 (Rat skeletal muscle myoblast) whole cell lysates at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 68 kDa

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• WB

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Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

All lanes:

Western blot - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (ab109025) at 1/1000 dilution

Lane 1:

NIH/3T3 cell lysate at 10 µg

Lane 2:

HaCaT cell lysate at 10 µg

Lane 3:

Neuro 2a cell lysate at 10 µg

Lane 4:

C2C12 cell lysate at 10 µg

Lane 5:

A431 cell lysate at 10 µg

Lane 6:

L6 cell lysate at 10 µg

Predicted band size: 68 kDa

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• OI-RD Scanning

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OI-RD Scanning - Anti-COX1 / Cyclooxygenase 1 antibody [EPR5866] (AB109025)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.