Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556]

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Mouse PBMC (mouse peripheral blood mononuclear cell) cells labelling 5 Lipoxygenase/5-LO with ab327006 at 1/500 (1.02 μg/ml) dilution , followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).

Confocal image showing nuclear staining in CD3 negative mouse PBMC (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Anti-mouse CD3 rat monoclonal antibody (Alexa Fluor® 647) was used to counterstain tubulin at 1/100 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunohistochemical analysis of paraffin-embedded (A) : Raw 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) cell pellet (B) : J774.1 (mouse reticulum cell sarcoma monocyte/macrophage) cell pellet (C) : NIH/3T3 (mouse embryonic fibroblast) cell pellets tissue labeling 5 Lipoxygenase/5-LO with ab327006 at 1/1000 (0.51 μg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) : Raw 264.7 cell pellet and (B) : J774.1 cell pellet, no staining on (C) : NIH/3T3 cell pellets. The primary antibody was incubated for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling 5 Lipoxygenase/5-LO with ab327006 at 1/4000 (0.128 μg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse spleen. The primary antibody was incubated for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background.

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunohistochemical analysis of paraffin-embedded Mouse skin tissue labeling 5 Lipoxygenase/5-LO with ab327006 at 1/4000 (0.128 μg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on immune cells in mouse skin. The primary antibody was incubated for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background.

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunohistochemical analysis of paraffin-embedded Mouse cardiac muscle tissue labeling 5 Lipoxygenase/5-LO with ab327006 at 1/4000 (0.128 μg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression : positive on scattered immune cells in mouse cardiac muscle. The primary antibody was incubated for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background.

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunohistochemical analysis of paraffin-embedded Mouse lung tissue labeling 5 Lipoxygenase/5-LO with ab327006 at 1/4000 (0.128 μg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on immune cells in mouse lung. The primary antibody was incubated for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background.

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Mouse PBMC (mouse peripheral blood mononuclear cell) cells labelling 5 Lipoxygenase/5-LO with ab327006 at 1/500 (1.02 μg/ml) dilution , followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).

Confocal image showing nuclear staining in CD11b positive mouse PBMC (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab8878 Anti-CD11b Rat monoclonal antibody was used to counterstain tubulin at 1/100 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunohistochemical analysis of paraffin-embedded Mouse lung carcinoma tissue labeling 5 Lipoxygenase/5-LO with ab327006 at 1/4000 (0.128 μg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on immune cells in mouse lung carcinoma. The primary antibody was incubated for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background.

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized J774A.1 (mouse reticulum cell sarcoma monocyte/macrophage) NIH/3T3 (mouse embryonic fibroblast) cells labelling 5 Lipoxygenase/5-LO with ab327006 at 1/500 (1.02 μg/ml) dilution , followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).

Confocal image showing nuclear staining in J774A.1 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). Low expression : NIH/3T3

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.

• WB

Lab

Western blot - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

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

Low expression : NIH/3T3.

The identity of the band higher than 250 kDa is unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

All lanes:

Western blot - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (ab327006) at 1/1000 dilution

Lane 1:

RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 2:

J774A.1 (mouse reticulum cell sarcoma monocyte/macrophage) whole cell lysate at 20 µg

Lane 3:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 77 kDa,36 kDa

false

Exposure time: 3s

• WB

Lab

Western blot - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (AB327006)

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

Low expression : heart.

The identity of the bands higher than 150 kDa are unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

All lanes:

Western blot - Anti-5 Lipoxygenase/5-LO antibody [EPR30234-556] (ab327006) at 1/1000 dilution

Lane 1:

Mouse lung tissue lysate at 20 µg

Lane 2:

Mouse spleen tissue lysate at 20 µg

Lane 3:

Mouse skin tissue lysate at 20 µg

Lane 4:

Mouse colon tissue lysate at 20 µg

Lane 5:

Mouse heart tissue lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 77 kDa,36 kDa

false

Exposure time: 10s