All lanes : Anti-RBX1 antibody [EPR20185] (ab221548) at 1/5000 dilution

Lane 1 : HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : HepG2 (human liver hepatocellular carcinoma cell line) whole cell lysate
Lane 3 : HT1080 (human fibrosarcoma cell line) whole cell lysate
Lane 4 : HEK-293 (human epithelial cell line from embryonic kidney) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Developed using the ECL technique.

Predicted band size: 12 kDa
Observed band size: 11,12 kDa why is the actual band size different from the predicted?

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Blocking and dilution buffer: 5% NFDM/TBST.

Exposure time: Lanes 1-3: 30 seconds; Lane 4: 10 seconds.

The molecular weight observed is consistent with what has been described in the literature (PMID: 22822056).

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded human colon tissue labeling RBX1 with ab221548 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear staining is observed on human colon tissue section. As documented in the literature RBX1 has lower expression level in normal tissue compared to cancerous tissue (PMID:19509229). Counter stained with hematoxylin. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling RBX1 with ab221548 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear and weak cytoplasmic staining on HeLa cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

All lanes : Anti-RBX1 antibody [EPR20185] (ab221548) at 1/1000 dilution

Lane 1 : Human fetal heart lysate
Lane 2 : Human fetal kidney lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/4000 dilution

Developed using the ECL technique.

Predicted band size: 12 kDa
Observed band size: 12 kDa


Exposure time: 15 seconds

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Blocking and dilution buffer: 5% NFDM/TBST.

All lanes : Anti-RBX1 antibody [EPR20185] (ab221548) at 1/1000 dilution

Lane 1 : Mouse heart lysate
Lane 2 : Mouse kidney lysate
Lane 3 : Mouse spleen lysate
Lane 4 : Rat brain lysate
Lane 5 : Rat kidney lysate
Lane 6 : Rat spleen lysate
Lane 7 : RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
Lane 8 : PC-12 (rat adrenal gland pheochromocytoma cell line) whole cell lysate
Lane 9 : NIH/3T3 (mouse embyro fibroblast cell line) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Developed using the ECL technique.

Predicted band size: 12 kDa
Observed band size: 11,12 kDa why is the actual band size different from the predicted?

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Exposure time: Lanes 1-6: 3 seconds; Lanes 7-8: 5 seconds.

Blocking and dilution buffer: 5% NFDM/TBST.

The molecular weight observed is consistent with what has been described in the literature (PMID: 22822056).

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded human colon carcinoma tissue labeling RBX1 with ab221548 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear and cytoplasmic staining is observed on human colon carcinoma tissue sections. As documented in the literature RBX1 has higher expression level in cancerous tissue compared to normal tissue (PMID:19509229). Counter stained with hematoxylin. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded human lung and lung carcinoma tissues labeling RBX1 with ab221548 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Weak nuclear staining of RBX1 on the epithelium cells of human lung (left) compared to strong staining in human lung carcinoma (right). As documented in the literature ROC1 has higher expression level in cancerous tissue compared to normal tissue (PMID:19509229). Counter stained with hematoxylin. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded human gastric carcinoma tissue labeling RBX1 with ab221548 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear and cytoplasmic staining on human gastric carcinoma tissue sections. The staining pattern observed is consistent with what has been described in the literature (PMID:24292229). Counter stained with hematoxylin. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded human bladder cancer tissue labeling RBX1 with ab221548 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear and cytoplasmic staining on human bladder carcinoma tissue sections. The staining pattern observed is consistent with what has been described in the literature (PMID:23667514). Counter stained with hematoxylin. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded mouse stomach tissue labeling RBX1 with ab221548 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear and cytoplasmic staining on mouse stomach tissue sections. Counter stained with hematoxylin. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of paraffin-embedded rat colon tissue labeling RBX1 with ab221548 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear and cytoplasmic staining on rat colon tissue sections. Counter stained with hematoxylin. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embyro fibroblast cell line) cells labeling RBX1 with ab221548 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear and weak cytoplasmic staining on NIH/3T3 cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution. Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cell line labeling RBX1 with ab221548 at 1/600 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized NIH/3T3 (mouse embyro fibroblast cell line) cell line labeling RBX1 with ab221548 at 1/60 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

This data was developed using ab221548, the same antibody clone in a different buffer formulation.RBX1 was immunoprecipitated from 0.35 mg of HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab221548 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab221548 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution. Lane 1: HeLa whole cell lysate 10 µg (Input). Lane 2: ab221548 IP in HeLa whole cell lysate. Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab221548 in HeLa whole cell lysate. Blocking and dilution buffer and concentration: 5% NFDM/TBST. Exposure time: 30 seconds. The molecular weight observed is consistent with what has been described in the literature (PMID: 22822056).