Anti-Heme Oxygenase 1 antibody [EPR18161-128]

• WB

Lab

Western blot - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

False colour image of Western blot : Anti-Heme Oxygenase 1 antibody [EPR18161-128] staining at 1/2000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab189491 was shown to bind specifically to Heme Oxygenase 1. A band was observed at 32 kDa in wild-type A549 cell lysates with no signal observed at this size in HMOX1 knockout cell line ab269503 (HMOX knockout A549 lysate ab259782). To generate this image, wild-type and HMOX1 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 5 % 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-Heme Oxygenase 1 antibody [EPR18161-128] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/heme-oxygenase-1-antibody-epr18161-128-bsa-and-azide-free-ab224677'>ab224677</a>) at 1/2000 dilution

Lanes 1 - 5:

Western blot - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (ab189491) at 1/2000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

HMOX1 knockout A549 cell lysate at 20 µg

Lane 3:

Mouse Spleen cell lysate at 20 µg

Lane 4:

HeLa cell lysate at 20 µg

Lane 5:

HEK-293 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: 32 kDa

false

• WB

Lab

Western blot - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Blocking : 5% NFDM/TBST.

Exposure time :

Lanes 1,2 and 3 : 2 seconds;

Lane 4 : 1 second

The molecular weight observed is consistent with what has been described in the literature (PMID : 18400743).

The lower band observed is a truncated form of Heme Oxygenase 1 (PMID : 17430897).

All lanes:

Western blot - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (ab189491) at 1/5000 dilution

Lane 1:

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

Lane 2:

Rat spleen lysate at 20 µg

Lane 3:

HeLa (human cervix adenocarcinoma epithelial cell) lysate at 20 µg

Lane 4:

Mouse spleen lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 33 kDa

Observed band size: 28 kDa,32 kDa

false

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Immunofluorescent analysis of 4% paraformaldehyde fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labeling Heme Oxygenase 1 with ab189491 at 1/250 dilution, followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on NIH/3T3 cells. Details of counterstains : ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at a 1/200 dilution; DAPI for nuclei.
The negative controls are as follows : Secondary antibody only for control.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Intracellular flow cytometric analysis of 90% methanol/ 4% paraformaldehyde fixed NIH/3T3 (mouse embryonic fibroblast) cell line labeling Heme Oxygenase1 with ab189491 at 1/50 (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluorr^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Intracellular flow cytometric analysis of 90% methanol/4% paraformaldehyde fixed HeLa (human cervix adenocarcinoma epithelial cell) cell line labeling Heme Oxygenase1 with ab189491 at 1/50 (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluorr^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Immunohistochemical analysis of paraffin embedded rat liver tissue labeling Heme Oxygenase 1 with ab189491 at 1/20,000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Positive staining on Kupffer cells of rat liver (PMID : 9449694) is observed. Counter stained with hematoxylin.
Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Immunohistochemical analysis of paraffin embedded human liver tissue labeling Heme Oxygenase 1 with ab189491 at 1/20,000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Positive staining on Kupffer cells of human liver (PMID : 9449694) is observed. Counter stained with hematoxylin.
Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Immunofluorescent analysis of 4% paraformaldehyde fixed, 0.1% Triton X-100 permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labeling Heme Oxygenase 1 with ab189491 at 1/250 dilution, followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on HeLa cells. Details of counterstains : ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at a 1/200 dilution; DAPI for nuclei.
The negative controls are as follows : Secondary antibody only for control.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Immunohistochemical analysis of paraffin embedded mouse liver tissue labeling Heme Oxygenase 1 with ab189491 at 1/20000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Positive staining on Kupffer cells of mouse liver (PMID : 9449694) is observed. Counter stained with hematoxylin.
Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

• IP

Lab

Immunoprecipitation - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Heme Oxygenase 1 was immunoprecipitated from 0.35 mg of NIH/3T3 (mouse embryonic fibroblast) whole cell lysate with ab189491 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab189491 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.
Lane 1 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate 10 μg (Input).
Lane 2 : NIH/3T3 whole cell lysate (+).
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab189491 in NIH/3T3 whole cell lysate (-).
Blocking and dilution buffer and concentration : 5% NFDM/TBST.

The truncated form of Heme Oxygenase 1 is described in the literature (PMID : 17430897).

All lanes:

Immunoprecipitation - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (ab189491)

Predicted band size: 33 kDa

Observed band size: 28 kDa,32 kDa

false

Exposure time: 1s

• WB

Lab

Western blot - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (AB189491)

Blocking : 5% NFDM/TBST.

The molecular weight observed is consistent with what has been described in the literature (PMID : 18400743).

The lower band observed is a truncated form of Heme Oxygenase 1 (PMID : 17430897).

All lanes:

Western blot - Anti-Heme Oxygenase 1 antibody [EPR18161-128] (ab189491) at 1/2000 dilution

All lanes:

Human spleen lysate at 10 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/4000 dilution

Predicted band size: 33 kDa

Observed band size: 28 kDa,32 kDa

false

Exposure time: 3min