Anti-AKR1C3+AKR1C4 antibody [EPR16726]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (AB209899)

Immunofluorescent analysis of 4% paraformaldehyde-fixed 0.1% Triton X-100 permeabilized A549 (Human lung carcinoma cell line) cells labeling AKR1C3 with ab209899 at 1/100 dilution followed by Goat anti-Rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing cytoplasmic staining on A549 cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG (Alexa Fluor^®594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows : -

-ve control 1 : ab209899 at 1/100 dilution followed by ab150120 at 1/1000 dilution.

-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (AB209899)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed A549 (Human lung carcinoma cell line) cells labeling AKR1C3with ab209899 at 1/70 dilution (red) compared with a Rabbit IgG,monoclonal- Isotype control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti Rabbit IgG (Alexa Fluor^® 488) at 1/500 dilution was used as the secondary antibody.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (AB209899)

Immunofluorescent analysis of 4% paraformaldehyde-fixed 0.1% Triton X-100 permeabilized HepG2 (Human liver hepatocellular carcinoma cell line) cells labeling AKR1C3 with ab209899 at 1/100 dilution followed by Goat Anti-Rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing cytoplasmic staining on HepG2 cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG (AlexaFluor^®594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows : -

-ve control 1 : ab209899 at 1/100 dilution followed by ab150120 at 1/1000 dilution.

-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (AB209899)

AKR1C3 was immunoprecipitated from 1mg of HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate with ab209899 at 1/30 dilution.

Western blot was performed from the immunoprecipitate using ab209899 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : HepG2 whole cell lysate, 10μg (Input).

Lane 2 : ab209899 IP in HepG2 whole cell lysate.

Lane 3 : Rabbit IgG,monoclonal - Isotype Control (ab172730) instead of ab209899 in HepG2 whole cell lysate.

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

Exposure time : 1 second.

All lanes:

Immunoprecipitation - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (ab209899)

Predicted band size: 37 kDa

false

• WB

Supplier Data

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (AB209899)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (ab209899) at 1/2000 dilution

Lane 1:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate (control) at 10 µg

Lane 2:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 10 µg with AKR1C3 peptide on the membrane

Lane 3:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 10 µg with AKR1C4 peptide on the membrane

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: 37 kDa

Observed band size: 37 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (AB209899)

Blocking and dilution buffer : 5% NFDM/TBST

Lanes 1 - 4:

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (ab209899) at 1/1000 dilution

Lanes 1 - 4:

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/akr1c3akr1c4-antibody-epr16726-bsa-and-azide-free-ab251511'>ab251511</a>) at 1/1000 dilution

Lane 1:

His-tagged human AKR1C1 recombinant protein, full-length

Lane 2:

His-tagged human AKR1C2 recombinant protein, full-length

Lane 3:

His-tagged human AKR1C3 recombinant protein, full-length

Lane 4:

His-tagged human AKR1C4 recombinant protein, full-length

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/20000 dilution

Observed band size: 39 kDa

false

Exposure time: 10s

• WB

Supplier Data

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (AB209899)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1/3 : 2 seconds; Lane 2 : 30 seconds.

Lanes 1 - 2:

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (ab209899) at 1/1000 dilution

Lane 3:

Western blot - Anti-AKR1C3+AKR1C4 antibody [EPR16726] (ab209899) at 1/5000 dilution

Lane 1:

Human fetal kidney lysate at 20 µg

Lane 2:

MOLT-4 (Human lymphoblastic leukemia cell line) whole cell lysate at 20 µg

Lane 3:

A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 37 kDa,90 kDa

Observed band size: 37 kDa

false