Anti-CRBN antibody [EPR28739-64]

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-CRBN antibody [EPR28739-64] (AB315344)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (human cervical adenocarcinoma epithelial cell) cells labelling CRBN with ab315344 at 1/500 dilution (0.1 ug)/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, ab150081) at 1/5000 dilution was used as the secondary antibody.

• IP

Supplier Data

Immunoprecipitation - Anti-CRBN antibody [EPR28739-64] (AB315344)

CRBN was immunoprecipitated from 0.35 mg HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate with ab315344 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315344 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

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

All lanes:

Immunoprecipitation - Anti-CRBN antibody [EPR28739-64] (ab315344) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate

Lane 2:

ab315344 at 1/30 IP in 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 ab315344 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

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-CRBN antibody [EPR28739-64] (AB315344)

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

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

All lanes:

Western blot - Anti-CRBN antibody [EPR28739-64] (ab315344) at 1/1000 dilution

Lane 1:

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

Lane 2:

HeLa transfected with siRNA specifically targeting CRBN 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

Observed band size: 50 kDa,36 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-CRBN antibody [EPR28739-64] (AB315344)

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

The molecular weight observed around 100 kDa is autoubiquitination of CRBN, which is consistent with what has been described in the literature (PMID : 25043012).

Lanes 1-3 are applied with Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 and lanes 4-9 are applied with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-CRBN antibody [EPR28739-64] (ab315344) at 1/1000 dilution

Lane 1:

Human hippocampus tissue lysate at 20 µg

Lane 2:

Human testis tissue lysate at 20 µg

Lane 3:

Human skeletal muscle tissue lysate at 20 µg

Lane 4:

Mouse brain tissue lysate at 20 µg

Lane 5:

Mouse testis tissue lysate at 20 µg

Lane 6:

Mouse skeletal muscle tissue lysate at 20 µg

Lane 7:

Rat brain tissue lysate at 20 µg

Lane 8:

Rat testis tissue lysate at 20 µg

Lane 9:

Rat skeletal muscle tissue lysate at 20 µg

Secondary

Lanes 1 - 3:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG (Merck DC03L) at 1/100000 dilution

Lanes 4 - 9:

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 1/2000 dilution

Observed band size: 50 kDa,36 kDa

false

Exposure time: 3s

• WB

Supplier Data

Western blot - Anti-CRBN antibody [EPR28739-64] (AB315344)

The molecular weight observed around 100 kDa is autoubiquitination of CRBN, which is consistent with what has been described in the literature (PMID : 25043012).

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

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-CRBN antibody [EPR28739-64] (ab315344) at 1/1000 dilution

Lane 1:

MOLT-4 (human lymphoblastic leukemia T lymphoblast) whole cell lysate at 40 µg

Lane 2:

HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Lane 3:

U266B1 (human mtiple myeloma B lymphocyte) whole cell lysate at 40 µg

Lane 4:

PC-3 (human prostate adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Lane 5:

HEK-293 (human embryonic kidney epithelial cell) transfected with scrambled siRNA control whole cell lysate at 40 µg

Lane 6:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 40 µg

Lane 7:

RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 40 µg

Lane 8:

C6 (rat glial tumor glial cell) whole cell lysate at 40 µg

Lane 9:

PC-12 (rat adrenal gland pheochromocytoma cell) whole cell lysate at 40 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

Observed band size: 50 kDa,36 kDa

false

Exposure time: 5.5s