Anti-HSPC150/UBE2T antibody [EP9447] - BSA and Azide free

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-HSPC150/UBE2T antibody [EP9447] - BSA and Azide free (AB284829)

This data was developed using ab179802, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa cells labelling HSPC150/UBE2T with ab179802 at 1/50 (2 ug/ml) dilution, followed by AlexaFluor^® 488-conjugated Goat anti-Rabbit secondary antibody (ab150081) at 1/1000 dilution at RT for 45 min. (Green).

Confocal image showing nuclear staining in HeLa cells (shown in green). ab195889 AlexaFluor^® 594-conjugated Anti-alpha Tubulin, used as a counterstain at 1/200 dilution, was co-incubated with ab179802 overnight at 4° C. Nuclear DNA was labeled with DAPI (shown in blue).

Secondary antibody only control : Secondary antibody is AlexaFluor^® 488-conjugated Goat anti-Rabbit secondary antibody (ab150081) at 1/1000 dilution.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-HSPC150/UBE2T antibody [EP9447] - BSA and Azide free (AB284829)

This data was developed using ab179802, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa cells labelling HSPC150/UBE2T with ab179802 at 1/20 dilution (5 ug/ml).

Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150081) was used as a secondary antibody.

• IP

Lab

Immunoprecipitation - Anti-HSPC150/UBE2T antibody [EP9447] - BSA and Azide free (AB284829)

This data was developed using ab179802, the same antibody clone in a different buffer formulation.

HSPC150/UBE2T was immunoprecipitated from 0.35 mg HeLa whole cell lysate with ab179802 at 1/20 dilution (0.5ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab179802 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used as secondary antibody at 1/5000 dilution.

All lanes:

Immunoprecipitation - Anti-HSPC150/UBE2T antibody [EP9447] (<a href='/en-us/products/primary-antibodies/hspc150-ube2t-antibody-ep9447-ab179802'>ab179802</a>) at 1/1000 dilution

Lane 1:

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

Lane 2:

HeLa 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/hspc150-ube2t-antibody-ep9447-ab179802'>ab179802</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: 23 kDa

false

Exposure time: 15s

• WB

Lab

Western blot - Anti-HSPC150/UBE2T antibody [EP9447] - BSA and Azide free (AB284829)

This data was developed using ab179802, the same antibody clone in a different buffer formulation.

An approximately 19 kDa band is observed but we are not sure how do define it

ab181602 was used as a GAPDH loading control.

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

All lanes:

Western blot - Anti-HSPC150/UBE2T antibody [EP9447] (<a href='/en-us/products/primary-antibodies/hspc150-ube2t-antibody-ep9447-ab179802'>ab179802</a>) at 1/10000 dilution

Lane 1:

A375 (Human malignant melanoma epithelial cell) whole cell lysate at 15 µg

Lane 2:

U-87 MG (Human glioblastoma-astrocytoma epithelial cell) whole cell lysate at 15 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: 23 kDa

true

Exposure time: 180s