Ab209665 staining Brachyury/Bry in MUG-Chor1 (human sacral bone chordoma) cells by ICC/IF (Immunocytochemistry/Immunofluorescence). Cells were fixed 4% paraformaldehyde and permeabilized with 0.1% tritonX-100. Samples were incubated with primary antibody at a 1/1000 dilution.  An AlexaFluor^®488 Goat anti-Rabbit (ab150077) was used as the secondary antibody at a 1/1000 dilution.  An Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) was used as a counterstain antibody.  DAPI was used as nuclear counterstain. Nuclear staining on MUG-Chor1 cells.

ab209665 staining Brachyury/Bry in mouse E14.5 embryo tissue sections by Immunohistochemistry (IHC-P). Tissue was fixed paraffin and antigen retrieval was performed by heat mediation using ab93684 (Tris/EDTA buffer, pH 9.0). Samples were incubated with primary antibody at a 1/4000 dilution. A ready to use Goat anti-rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counter-stain. Nuclear staining on notochord of mouse E14.5 embryo.

Anti-Brachyury / Bry antibody [EPR18113] (ab209665) at 1/1000 dilution + MUG-Chor1 (human sacral bone chordoma), whole cell lysate at 10 µg

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

Predicted band size: 47 kDa
Observed band size: 43,49 kDa why is the actual band size different from the predicted?


Exposure time: 5 seconds

Blocking and diluting buffer: 5% NFDM/TBST

Ab209665 staining Brachyury/Bry in MUG-Chor1 (Human sacral bone chordoma) cell line by Flow cytometry. Cells were fixed with 4% paraformaldehyde and 90% methanol. Sample was incubated with primary antibody at 1/400 dilution (red). A Goat anti-rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution.  Rabbit monoclonal IgG (ab172730) (Black) was used as an isotype control. Cell without incubation with primary antibody and secondary antibody (Blue).

Lane 1 (input):  MUG-Chor1 (human sacral bone chordoma) whole cell lysate, 10 μg
Lane 2 (+): MUG-Chor1 whole cell lysate
Lane 3 (-):  Rabbit monoclonal IgG (ab172730) instead of  ab209665 in MUG-Chor1 whole cell lysate

Ab209665 immunoprecipitating Brachyury/Bry in MUG-Chor1 lysates. For western blotting, primary antibody used was ab209665 at 1/1000 dilution.  VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution. Blocking and diluting buffer used was 5% NFDM/TBST.

Ab209665 staining Brachyury/Bry in Rat E14.5 embryo tissue sections by Immunohistochemistry (IHC-P). Tissue was fixed paraffin and antigen retrieval was performed by heat mediation using ab93684 (Tris/EDTA buffer, pH 9.0). Samples were incubated with primary antibody at a 1/4000 dilution. A ready to use Goat anti-rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counter-stain. Nuclear staining on notochord of rat E14.5 embryo.

IHC - Wholemount of E6.5 wholemount mouse embryo labelling Brachyury / Bry with ab209665. Sample was incubated with primary antibody (1/100 in PBS/0.1% Triton-X/5% donkey serum/1% BSA) for 18 hours at 4°C. An Alexa Fluor^® 488-conjugated donkey anti-rabbit IgG monoclonal was used as the secondary antibody (1/500).

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Immunohistochemical analysis of paraffin-embedded Human chordoma tissue labeling Brachyury / Bry with ab209665 at 1/8000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nucleus staining on Human chordoma 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) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling Brachyury / Bry with ab209665 at 1/8000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative staining on Human kidney. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Human meningioma tissue labeling Brachyury / Bry with ab209665 at 1/8000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative staining on Human meningioma. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Human chondrosarcoma tissue labeling Brachyury / Bry with ab209665 at 1/8000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative staining on Human chondrosarcoma. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

 

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.