Anti-RBPJK antibody [EPR13479]

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-RBPJK antibody [EPR13479] (AB180588)

Immunocytochemistry/ Immunofluorescence analysis of MG 63 (human osteosarcoma fibroblast) cells labeling RBPJK with Purified ab180588 at 1 : 50 dilution. Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) (ab195889) at 1 : 200 dilution. Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150081) was used as the secondary antibody at 1 : 1000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control.

Confocal image showing mainly nuclear staining in MG-63 cell line(shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).

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

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-RBPJK antibody [EPR13479] (AB180588)

Flow cytometry overlay histogram of 4% Paraformaldehyde MG 63 (human osteosarcoma fibroblast) cells permeabilised with 90% methanol labelling RBPJK with ab180588 at 1/50 dilution (red) compared with a Rabbit monoclonal IgG (ab172730) (Black) and an unlabeled 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.

• WB

Supplier Data

Western blot - Anti-RBPJK antibody [EPR13479] (AB180588)

All lanes:

Western blot - Anti-RBPJK antibody [EPR13479] (ab180588) at 1/2000 dilution

Lane 1:

293 cell line lysate

Lane 2:

MCF7 cell line lysate

Lane 3:

HeLa cell line lysate

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 56 kDa

Observed band size: 60 kDa

false

• WB

Lab

Western blot - Anti-RBPJK antibody [EPR13479] (AB180588)

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

Extra band at : 90kDa
We are unsure as to the identity of this extra band.

For Lysate preparation protocol, please refer to the protocol section of the website and/or here.

All lanes:

Western blot - Anti-RBPJK antibody [EPR13479] (ab180588) at 1/1000 dilution

Lane 1:

MCF-7 (Human breast adenocarcinoma epithelial cell) whole cell lysate 20ug prepared using RIPA lysis method at 20 µg

Lane 2:

MCF-7 (Human breast adenocarcinoma epithelial cell) whole cell lysate 20ug prepared using 1% SDS hot lysis method at 20 µg

Secondary

All lanes:

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

Predicted band size: 56 kDa

Observed band size: 60 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-RBPJK antibody [EPR13479] (AB180588)

The double bands are consistent with what has been described in the literature PMID : 9102301 and 24603501

All lanes:

Western blot - Anti-RBPJK antibody [EPR13479] (ab180588) at 1/1000 dilution

Lane 1:

Raji (Human Burkitt's lymphoma B lymphocyte) whole cell lysates at 15 µg

Lane 2:

MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates at 15 µg

Lane 3:

F9 (Mouse embryonal carcinoma epithelial cell) whole cell lysates at 15 µg

Lane 4:

C6 (Rat glial tumor glial cell) whole cell lysates 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: 56 kDa

Observed band size: 56-60 kDa

false

• WB

Lab

Western blot - Anti-RBPJK antibody [EPR13479] (AB180588)

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

All lanes:

Western blot - Anti-RBPJK antibody [EPR13479] (ab180588)

Lane 1:

THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Lane 2:

MG-63 (human osteosarcoma fibroblast) 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

Observed band size: 55 kDa

false

Exposure time: 48s

• WB

Lab

Western blot - Anti-RBPJK antibody [EPR13479] (AB180588)

Blocking and diluting buffer : 5% NFDM/TBST.

Exposure time : Lane 1-3 : 3 minutes
Lane 4 : 45 seconds

All lanes:

Western blot - Anti-RBPJK antibody [EPR13479] (ab180588) at 0.4 µg/mL

Lane 1:

C6 (Rat glial tumor glial cell) whole cell lysate at 20 µg

Lane 2:

PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

Raw264.7 (Mouse abelson murine leukemia virus-induced tumor macrophage) 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 1/50000 dilution

Predicted band size: 56 kDa

false