ENCAB282XQE

Alternate accession: ENCAB805FRK

Antibody against Homo sapiens EHMT2

Homo sapiens
HepG2, endothelial cell of umbilical vein, K562, GM12878, A549
characterized to standards with exemption
Status
released
Source (vendor)
Bethyl Labs
Product ID
A300-933A
Lot ID
1
Characterized targets
EHMT2 (Homo sapiens)
Host
rabbit
Clonality
polyclonal
Antigen description
H3K9 methyltransferase G9a
Aliases
bradley-bernstein:PchAb 255, michael-snyder:AS-1566
External resources

Characterizations

EHMT2 (Homo sapiens)
Method: ChIP-seq comparison
exempt from standards
Caption
This validation relies on the use of antibodies to a histone lysine methyltransferase (EHMT2-G9a) and a chromatin regulator (CBX3) in HepG2 cells, and the demonstration that highly similar patterns of enrichment are obtained with each antibody. The first track shown used an antibody to EHMT2 (PchAb 255, ENCAB282XQE), and the second track shown used an antibody to CBX3 submitted for characterization to Encode standards (PchAb 362, ENCAB000AHR).
Submitter comment
SAV7
Reviewer comment
Not compliant due to low genome-wide correlation (Correlation at 0.7147). Must be 0.8 or greater. However, this decision was overridden by Dr. Peggy Farnham (chair of the ENCODE antibody review committee) for the following reason ".. they were comparing datasets from EHMT2 and CBX3. Although I would expect the patterns to be quite similar, they will NOT be identical as CBX3 will not necessarily bind to all EHMT2 positions. Based on the visual inspection, the tracks (in combination with the primary characterization) do support the conclusion that the EHMT2 antibody is most likely correct. Therefore, assuming the datasets passed IDR, I would approve release with the same statement as before."
Submitted by
Nina Farrell
Lab
Bradley Bernstein, Broad
EHMT2 (Homo sapiens)
HepG2GM12878endothelial cell of umbilical veinA549
Method: immunoblot
Attachment from submitter
compliant
Caption
Nuclear lysates from K562 (10ug), HepG2 (10ug), GM12878 (7ug), HUVEC (10ug), were loaded into a 4-12% Bis-Tris gel in 1X MES running buffer. After separation, the samples were transferred to a nitrocellulose membrane using the iblot system. Membrane was blocked for an hour in room temperature, with 5% BSA in TBS-T and blotted with primary antibody in the appropriate concentration over night at 4c. Membrane was washed and blotted with secondary HRP-conjugated antibody. Detection was made with Optiblot ECL Detect Kit (ab133406) for 2 min. A single band around the expected size was detected.
Submitter comment
The antibody review board exempted A549 data from primary characterization requirement based on the quality of the ChIP of the two replicates and the other cell lines 03-05-2018
Reviewer comment
The expected size is ~132 kDa but runs near 165 kDa, which corresponds to the 165 kDa G9a-L isoform.
Submitted by
Noam Shoresh
Lab
Bradley Bernstein, Broad
EHMT2 (Homo sapiens)
K562
Method: immunoprecipitation
Attachment from submitter
compliant
Caption
Immunoprecipitation was performed on nuclear extracts from the cell line: K562, using the antibody A300-933A. The blot shows western blot analysis of input, flowthrough, immunoprecipitate and mock immunoprecipitate using IgG.Molecular Weight: 132.370
Reviewer comment
The expected size is ~132 kDa but runs near 165 kDa, which corresponds to the 165 kDa G9a-L isoform.
Submitted by
Denis Salins
Lab
Michael Snyder, Stanford