Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

Immunohistochemical analysis of formalin fixed paraffin embedded Human spleen tissue labelling eIF4A3 with ab180573 at 1/2000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Sections were counterstained with Hematoxylin.

Positive staining on human spleen.
The section was incubated with ab180573 at 4°C overnight.

Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH9.0)

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

Immunohistochemical analysis of paraffin-embedded Human prostate tissue labeling eIF4A3 with ab180573 at 1/100 dilution. Prediluted (ready to use) HRP Polymer for Rabbit IgG was used as a secondary antibody. Counter stain : Hematoxylin.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

Perform heat mediated antigen retrieval with EDTA buffer pH 9 before commencing with IHC staining protocol.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labelling eIF4A3 with ab180573 at 1/100 (9.39 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).

Nuclear DNA was labelled with DAPI (shown in blue). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 (2.5ug/ml) dilution (Red).

Confocal image showing nuclear staining in HeLa cells.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

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

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized MCF7 (human breast adenocarcinoma epithelial cell) cells labelling eIF4A3 with ab180573 at 1/90 dilution (1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

Goat anti rabbit IgG (Alexa Fluor^® 488, ab150097) at 1/5000 dilution was used as the secondary antibody.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

Immunofluorescent analysis of 4% paraformaldehyde MCF7 cells labeling eIF4A3 with ab180573 at 1/500 dilution (green), or with Dapi counter stain (blue). Goat anti rabbit IgG (Dylight 488) was used as a secondary antibody, at a dilution of 1/200.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

• IP

Lab

Immunoprecipitation - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

eIF4A3 was immunoprecipitated from 0.35 mg HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate with ab180573 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab180573 at 1/500 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate, 10 µg
Lane 2 : HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab180573 in HeLa whole cell lysate

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

All lanes:

Immunoprecipitation - Anti-eIF4A3 antibody [EPR14301(B)] (<a href='/en-us/products/primary-antibodies/eif4a3-antibody-epr14301b-ab180573'>ab180573</a>) at 1/500 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 10 µg

Lane 2:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of <a href='/en-us/products/primary-antibodies/eif4a3-antibody-epr14301b-ab180573'>ab180573</a> in HeLa whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Predicted band size: 47 kDa

false

Exposure time: 1s

• WB

Lab

Western blot - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

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

ab181602 was used as a GAPDH loading control.

ab180573 has a faint band above the target band, we recommend using ab180519 to get desire result in western blot.

All lanes:

Western blot - Anti-eIF4A3 antibody [EPR14301(B)] (<a href='/en-us/products/primary-antibodies/eif4a3-antibody-epr14301b-ab180573'>ab180573</a>) at 1/1000 dilution

Lane 1:

SK-OV-3 (Human ovarian cancer epithelial cell) whole cell lysate at 20 µg

Lane 2:

HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 5:

MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 6:

Raji (Human Burkitt's lymphoma B lymphocyte) whole cell lysate at 20 µg

Lane 7:

HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell 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 1/20000 dilution

Predicted band size: 47 kDa

Observed band size: 46 kDa

false

• WB

Lab

Western blot - Anti-eIF4A3 antibody [EPR14301(B)] - BSA and Azide free (AB236131)

Blocking and dilution buffer : 5% NFDM/TBST.

Anti-DDDDK tag (Binds to FLAG^® tag sequence) antibody (ab205606) used as loading control at 1/1000 dilution. The secondary antibody used is ab97051, a HRP goat anti-rabbit IgG (H+L), at a dilution of 1/20000.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab180573).

All lanes:

Western blot - Anti-eIF4A3 antibody [EPR14301(B)] (<a href='/en-us/products/primary-antibodies/eif4a3-antibody-epr14301b-ab180573'>ab180573</a>) at 1/1000 dilution

Lane 1:

eIF4A1 Human Recombinant Protein at 10 ng

Lane 2:

eIF4A2 Human Recombinant Protein at 10 ng

Lane 3:

eIF4A3 Human Recombinant Protein at 10 ng

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 1/20000 dilution

Predicted band size: 47 kDa

Observed band size: 46 kDa

false

Exposure time: 5s