ENCAB913HCF

Antibody against Homo sapiens EZH2

Homo sapiens
HeLa-S3, K562
characterized to standards
Homo sapiens
OCI-LY1, KMS-11, smooth muscle cell, GM23338, GM23248, SK-N-MC, A673, hepatocyte, fibroblast of arm, OCI-LY7, DOHH2, neural progenitor cell, HCT116
characterized to standards with exemption
Status
released
Source (vendor)
Cell Signaling
Product ID
5246S
Lot ID
6
Characterized targets
EZH2 (Homo sapiens)
Host
rabbit
Clonality
monoclonal
Antigen description
enhancer of zeste homolog 2 (Drosophila)
Aliases
bradley-bernstein:PchAb 1206, bradley-bernstein:PchAb 1290
External resources

Characterizations

EZH2 (Homo sapiens)
smooth muscle cellGM23338SK-N-MCA673neural progenitor cellHCT116K562fibroblast of armKMS-11hepatocyteOCI-LY7DOHH2OCI-LY1GM23248
Method: immunoblot
Attachment from submitter
exempt from standards
Caption
Both ENCODE submitter and reviewer have agreed that this is a well-established antibody lot. Hence it does not require additional validation as standardized by the ENCODE Consortium.
Submitter comment
The lab requests that this is considered a well-documented antibody lot and therefore does not require additional validation.
Reviewer comment
Dr. Peggy J Farnham: I would support a “no additional validation needed” classification for this antibody.
Submitted by
Nina Farrell
Lab
Bradley Bernstein, Broad
Download
EZH2 (Homo sapiens)
Method: ChIP-seq comparison
compliant
Caption
This validation relies on the use of antibodies to a chromatin regulator and a functionally related histone modification in A673 cells, and the demonstration that highly similar (>0.9) patterns of enrichment are obtained with each antibody. The first antibody is to phosphorylated EZH2 (PchAb 1206, ENCAB913HCF), and the second antibody is to H3K27me3 previously characterized to Encode standards (PchAb 848, ENCAB036YAO).
Submitted by
Nina Farrell
Lab
Bradley Bernstein, Broad
EZH2 (Homo sapiens)
Method: ChIP-seq comparison
compliant
Caption
This validation relies on the use of antibodies to a chromatin regulator and a functionally related histone modification in A673 cells, and the demonstration that highly similar (>0.9) patterns of enrichment are obtained with each antibody. The first antibody is to phosphorylated EZH2 (PchAb 1206, ENCAB913HCF), and the second antibody is to H3K27me3 previously characterized to Encode standards (PchAb 848, ENCAB036YAO).
Submitted by
Nina Farrell
Lab
Bradley Bernstein, Broad
EZH2 (Homo sapiens)
HeLa-S3K562
Method: immunoblot
compliant
Caption
We performed a WB using 0.7ug unmodified recombinant Histone H3 (Millipore 14-494), 20ug of HeLa nuclear lysate and 20ug of K562 nuclear lysate. Lysates were resolved by electrophoresis on a 4-12% bis-tris gel. After separation, the samples were transferred to a nitrocellulose membrane with an Invitrogen iBlot system. Membrane was blocked for an hour at room temperature, with 5% nonfat dry milk in TBST. The primary antibody was diluted in TBST and 5% BSA at the appropriate concentration over night at 4C. Membrane was washed and blotted with secondary HRP-conjugated antibody for 1 hour at room temp. Detection was made with Optiblot ECL Detect Kit (ab133406) for 1 min. Band of expected size (~98kDa) visualized in both nuclear lysates and could not be detected in the unmodified recombinant Histone H3. Antibody seems to be specific since no reactivity with non histone proteins or unmodified histones was detected.
Submitted by
Nina Farrell
Lab
Bradley Bernstein, Broad
EZH2 (Homo sapiens)
K562
Method: immunoprecipitation
Attachment from submitter
compliant
Caption
K562 whole cell lysate was immunoprecipitated using primary antibody, and the IP fraction along side a whole cell lysate were loaded on a CriterionXT gel and separated. After separation, the samples were transferred using a wet transfer. Blotting with primary (same as that used for IP) and secondary HRP-conjugated antibodies was performed. Band of expected size visualized representing strongest signal in the lane (IP-1).
Submitted by
Noam Shoresh
Lab
Bradley Bernstein, Broad