Caenorhabditis elegans
at least one cell type or tissue
not pursued
Drosophila melanogaster
at least one cell type or tissue
Homo sapiens
at least one cell type or tissue
Mus musculus
at least one cell type or tissue
Status
released
Source (vendor)
Millipore
Product ID
07-449
Lot ID
DAM1387952
Host
rabbit
Clonality
polyclonal
Purification
Protein A
Isotype
IgG
Antigen description
KLH-conjugated, synthetic 2X-branched peptide containing the sequence ...AR(me3K)SAP... in which me3K corresponds to trimethyl-lysine at residue 27 of human histone H3.

Characterizations

H3K27me3 (Homo sapiens)
Method: dot blot assay
Caption
Suitably modified peptides modeled on histone tails were obtained from Abcam. Each peptide is added to Bio-Dot apparatus at 0.01 and 0.1 ug / 50uL total volume (BioRad) Let sit on rocker 60 minutes.Pull through with vacuum, wash once with 100uL TBS and twice with 200uL TBSTw. Remove membrane from Bio-Dot and transfer to 10mL Pierce SuperBlock in tray. Place on rocker for 30 minutes. Dump off SuperBlock.Remove membrane from Bio-Dot and transfer to 10mL Pierce SuperBlock in tray.
Submitter comment
Lot mismatch: Image is for lot DAM1514011
Submitted by
Bradley Bernstein
Lab
Bradley Bernstein, Broad
H3K27me3 (Mus musculus)
Method: dot blot assay
Caption
Dr. Brad Bernstein and his colleagues at the Broad Institute have already validated this and several other antibodies directed against specific histone modifications. They spotted synthetic peptides containing one of about 20 histone modifications on a blot, in two different concentrations. The blot was then allowed to react with the antibody, and the antigen-antibody complexes were visualized and quantified. In each case, the antibody showed strong specificity. This is far better than showing a single band on a Western blot, since all the modifications we examine are on histone H3, and they all will show the H3 band. The Western blot will not demonstrate specificity for a particular modification, whereas the dot blot does. Our validation point for mouse is that there is nothing species-specific about the existing validations. Synthetic peptides were used on the blot, and the assay was for specific reaction with the antibody. The peptides on the blot were not species-specific because HUMAN AND MOUSE HISTONE H3 ARE IDENTICAL IN THE RELEVANT REGIONS. Human and mouse H3 differ at only one position, amino acid 97, where a Cys in human is replaced by a Ser in mouse. There are NO differences in the relevant region, which is the N-terminal 36 amino acids.
Submitter comment
Lot mismatch: Dot blot referred to is for lot DAM1514011.
Submitted by
Ross Hardison
Lab
Ross Hardison, PennState