Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free (AB303525)

This data was developed using ab303524, the same antibody clone in a different buffer formulation. Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized PC-3 (human prostate adenocarcinoma epithelial cell) cells labelling eRF3A/GSPT1 with ab303524 at 1/100 (4.94 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). Confocal image showing mainly cytoplasmic staining in PC-3 cell lines. The image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5 ug/ml dilution (Red). 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 2 ug/ml dilution.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free (AB303525)

This data was developed using ab303524, the same antibody clone in a different buffer formulation. Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized PC-3 (human prostate adenocarcinoma epithelial cell) cells labelling eRF3A/GSPT1 with ab303524 at 1/500 dilution (0.1ug) (Red) (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). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.

• IP

Supplier Data

Immunoprecipitation - Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free (AB303525)

This data was developed using ab303524, the same antibody clone in a different buffer formulation. eRF3A/GSPT1 was immunoprecipitated from 0.35 mg PC-3 (human prostate adenocarcinoma epithelial cell), whole cell lysate 10 ug with ab303524 at 1/30 dilution (2 µg in 0.35 mg lysates). Western blot was performed on the immunoprecipitate using ab303524 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : PC-3 (human prostate adenocarcinoma epithelial cell), whole cell lysate 10 µg (Inset) Lane 2 : ab303524 IP in PC-3 whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab303524 in PC-3 whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-eRF3A/GSPT1 antibody [EPR27296-4] (<a href='/en-us/products/primary-antibodies/erf3a-gspt1-antibody-epr27296-4-ab303524'>ab303524</a>) at 1/30 dilution

Lane 1:

PC-3 (human prostate adenocarcinoma epithelial cell) at 10 µg

Lane 2:

<a href='/en-us/products/primary-antibodies/erf3a-gspt1-antibody-epr27296-4-ab303524'>ab303524</a> IP in PC-3 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: 56 kDa

Observed band size: 80 kDa

false

Exposure time: 3s

• WB

Supplier Data

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free (AB303525)

This data was developed using ab303524, the same antibody clone in a different buffer formulation. Blocking / Diluting buffer and concentration : 5% NFDM/TBST

All lanes:

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] (<a href='/en-us/products/primary-antibodies/erf3a-gspt1-antibody-epr27296-4-ab303524'>ab303524</a>) at 1/1000 dilution

Lane 1:

HEK-293T transfected with human ERF3A expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 2:

HEK-293T transfected with human ERF3B expression vector containing a myc-His-tag®, 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 1/100000 dilution

Predicted band size: 56 kDa

Observed band size: 80 kDa

false

Exposure time: 5.5s

• WB

Supplier Data

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free (AB303525)

This data was developed using ab303524, the same antibody clone in a different buffer formulation. Blocking / Diluting buffer and concentration : 5% NFDM/TBST Exposure time : Lane 1 : 136 seconds Lane 2-5 : 48 seconds In lane 1, the lysates were stored at -80℃ prior to Western Blotting. The bands beneath the target band (80 kDa) are expected to be degradation products. In lane 2-5, the lysates were freshly made and used for Western Blotting immediately to minimize protein degradation.

All lanes:

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] (<a href='/en-us/products/primary-antibodies/erf3a-gspt1-antibody-epr27296-4-ab303524'>ab303524</a>) at 1/1000 dilution

Lanes 1 - 2:

HeLa (human epithelial cell line from cervical adenocarcinoma), whole cell lysate at 20 µg

Lane 3:

293T (human embryonic kidney epithelial cell), whole cell lysate at 20 µg

Lane 4:

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

Lane 5:

PC-12 (rat adrenal gland pheochromocytoma), 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 1/100000 dilution

Predicted band size: 56 kDa

Observed band size: 80 kDa

false

• WB

Supplier Data

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free (AB303525)

This data was developed using ab303524, the same antibody clone in a different buffer formulation.

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

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

All lanes:

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] (<a href='/en-us/products/primary-antibodies/erf3a-gspt1-antibody-epr27296-4-ab303524'>ab303524</a>) at 1/1000 dilution

All lanes:

Rat spleen tissue 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: 56 kDa

Observed band size: 80 kDa

true

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] - BSA and Azide free (AB303525)

This data was developed using ab303524, the same antibody clone in a different buffer formulation.

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

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

Lysates were freshly made and used immediately to minimize protein degradation.

All lanes:

Western blot - Anti-eRF3A/GSPT1 antibody [EPR27296-4] (<a href='/en-us/products/primary-antibodies/erf3a-gspt1-antibody-epr27296-4-ab303524'>ab303524</a>) at 1/1000 dilution

Lane 1:

Mouse brain tissue lysate at 20 µg

Lane 2:

Mouse testis tissue lysate at 20 µg

Lane 3:

Mouse spleen tissue 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: 56 kDa

Observed band size: 80 kDa

true

Exposure time: 3min