All lanes : Anti-RUNX3 antibody [EPR20680] (ab224642) at 1/1000 dilution

Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : RUNX3 knockout HAP1 whole cell lysate
Lane 3 : Ramos whole cell lysate
Lane 4 : HeLa whole cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 44, 46 kDa

This data was developed using ab224642, the same antibody clone in a different buffer formulation.

Lanes 1 - 4: Merged signal (red and green). Green - ab224642 observed at 44-46 kDa. Red - loading control, ab130007, observed at 130 kDa.

ab224642 was shown to recognize RUNX3 in wild-type HAP1 cells as signal was lost at the expected MW in RUNX3 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and RUNX3 knockout samples were subjected to SDS-PAGE. ab224642 and ab130007 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes : Anti-RUNX3 antibody [EPR20680] (ab224642) at 1/1000 dilution

Lane 1 : Raji (human Burkitt's lymphoma cell line) whole cell lysate
Lane 2 : Ramos (human Burkitt's lymphoma cell line) whole cell lysate
Lane 3 : MCF7 (human breast adenocarcinoma cell line) 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: 44, 46 kDa
Observed band size: 44,46 kDa why is the actual band size different from the predicted?


Exposure time: 10 seconds

This data was developed using ab224642, the same antibody clone in a different buffer formulation.

Blocking and dilution buffer: 5% NFDM/TBST.

Negative control: MCF-7 (PMID: 21706051).

This target detects both predicted isoforms 44KDa and 46KDa, consistent with what has been described in the literature (PMID: 23700080).

All lanes : Anti-RUNX3 antibody [EPR20680] (ab224642) at 1/1000 dilution

Lane 1 : Human thymus tissue lysate
Lane 2 : Human tonsil tissue 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: 44, 46 kDa
Observed band size: 44,46 kDa why is the actual band size different from the predicted?

This data was developed using ab224642, the same antibody clone in a different buffer formulation.

Blocking and dilution buffer: 5% NFDM/TBST.

Exposure times: Lane 1: 3 minutes; Lane 2: 15 seconds.

This target detects both predicted isoforms 44 kDa and 46 kDa, consistent with what has been described in the literature (PMID: 23700080).

This data was developed using ab224642, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of paraffin-embedded human stomach tissue labeling RUNX3 with ab224642 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear staining on lymphoid cells of human stomach (PMID:15514019; PMID:21786422) is observed. 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 ab224642, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of paraffin-embedded human gastric cancer tissue labeling RUNX3 with ab224642 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear staining on lymphoid cells of human gastric cancer (PMID:27566570). 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 ab224642, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of paraffin-embedded human diffuse large B cell lymphoma tissue labeling RUNX3 with ab224642 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Nuclear staining on human diffuse large B cell lymphoma (PMID:27184221). 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 ab224642, the same antibody clone in a different buffer formulation.RUNX3 was immunoprecipitated from 0.35 mg of Raji (human Burkitt's lymphoma cell line) whole cell lysate with ab224642 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab224642 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution. Lane 1: Raji whole cell lysate 10 µg (Input). Lane 2: ab224642 IP in Raji whole cell lysate. Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab224642 in Raji whole cell lysate. Blocking and dilution buffer and concentration: 5% NFDM/TBST. Exposure time: 10 second. The molecular weight observed is consistent with what has been described in the literature (PMID:23700080).