Anti-Brachyury / Bry antibody - Mesoderm Marker (ab20680)

Immunohistochemistry (Paraffin-embedded sections) - Brachyury / Bry antibody - Mesoderm Marker (ab20680)

A proportion of nuclei from Day 3 Mouse EBs (derived from R1 cells) stained positive for Brachury using ab20680. In Day 8 EBs a vastly reduced proportion of nuclei stained positive (not shown). In day 10 EBs no nuclei stained Brachyury-positive. The IHC-P data for Brachyury in EBs corresponded with qRT-PCR expression data (not shown). Controls on Day 3 EBs in which primary or secondary antibody were omitted, undifferentiated ES cells and feeder cells (MEFs) all exhibited very little background staining.

Zinc was used for tissue fixation.

Amanda Evans, University of Cambridge

Immunocytochemistry/ Immunofluorescence - Brachyury / Bry antibody - Mesoderm Marker (ab20680)

Anti-Brachyury antibody, ab20680, stained nuclei of human embryonic stem cells differentiated into mesendoderm. As would be expected, mesoderm cells express Brachyury whereas endoderm cells are negative.

Ludovic Valllier, University of Cambridge, UK

Western blot - Brachyury / Bry antibody - Mesoderm Marker (ab20680)

Anti-Brachyury / Bry antibody - Mesoderm Marker (ab20680) at 1 µg/ml + Murine ES Whole Cell Lysate Expressing Mesoderm Markers at 15 µg

Secondary
Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Performed under reducing conditions.

Predicted band size : 47 kDa
Observed band size : 53 kDa (why is the actual band size different from the predicted?)
Additional bands at : 75 kDa. We are unsure as to the identity of these extra bands.

ab20680 recognizes a band of 53 kDa in Murine Embryonic Stem Cells differentiated to express mesoderm markers. This corresponds in size to the Brachyury protein which has a predicted molecular weight of 47 kDa. A band of 75 kDa has also been observed, although we are unsure as to the identity of this band.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Brachyury / Bry antibody - Mesoderm Marker (ab20680)

ab20680 staining Brachyury in Human brain tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 3% serum for 30 minutes at room temperature; antigen retrieval was by heat mediation with a citrate buffer. Samples were incubated with primary antibody (1/2000 in horse serum) for 12 hours. A Streptavidin-conjugated Horse anti-rabbit monoclonal (1/250) was used as the secondary antibody.

This image is courtesy of an Abreview submitted by Jim Manavis