ENCAB000AMH

Antibody against Homo sapiens USF2, Mus musculus USF2

Homo sapiens
K562, GM12878, HeLa-S3, A549, IMR-90, SK-N-SH
characterized to standards with exemption
Homo sapiens
any cell type or tissue
partially characterized
Mus musculus
any cell type or tissue
Homo sapiens
HepG2
not characterized to standards
Status
released
Source (vendor)
Abcam
Product ID
ab60931
Lot ID
424648
Host
mouse
Clonality
monoclonal
Purification
Protein A/G
Antigen description
Immunogen corresponds to amino acids 1-101 of Human USF2.

Characterizations

USF2 (Homo sapiens)
A549IMR-90SK-N-SH
Method: immunoprecipitation
Attachment from submitter
exempt from standards
Caption
Primary characterizations were waived for this antibody in A549, IMR-90 and SK-N-SH on the basis of previous compliant primary and secondary characterizations in a different lot of this antibody (ENCAB483CQL).
Submitter comment
We ran out of this antibody lot and cannot perform characterizations in A549, IMR-90 and SK-N-SH but we have used a different lot of the same antibody to perform compliant characterizations (ENCAB483CQL).
Reviewer comment
The exemption request was granted by the Antibody Review Panel on May 28, 2017.
Submitted by
Jessika Adrian
Lab
Michael Snyder, Stanford
USF2 (Homo sapiens)
Method: knockdown or knockout
Attachment from submitter
exempt from standards
Caption
A new secondary characterization is not required as one exists for ENCAB940OHP which is a lot of the same antibody (ab60931). The primary characterization (https://www.encodeproject.org/antibody-characterizations/08379376-c82b-4a29-84ed-7d6f9a1adb21/) of K562 for this lot shows the same banding pattern as the primary characterization (https://www.encodeproject.org/antibody-characterizations/611d4051-bbdc-4862-8a7f-1c3532868920/) in K562 cell line for the lot ENCAB940OHP.
Submitter comment
This is a new lot of a previously characterized antibody ENCAB940OHP and we think no further characterization is necessary beyond the primary if the banding pattern is consistent with the old.
Reviewer comment
The banding in K562 is consistent between new and old lots so a new secondary characterization is not necessary according to the standards.
Submitted by
Jessika Adrian
Lab
Michael Snyder, Stanford
USF2 (Homo sapiens)
Method: immunoprecipitation
not reviewed
Caption
A major band of ~45kD is observed in all cell lines tested. This band is somewhat higher than expected for this protein (molecular weight of 37kD). However, this has been previously observed (see the image at http://www.abcam.com/USF2-antibody-ab60931.html) and may be due to post-translational modification. An additional band at ~100kD is sometimes seen in the K562 and HepG2 cells lines, but this band is not immunoprecipitated efficiently (panel B).
Submitted by
Michael Snyder
Lab
Michael Snyder, Stanford
USF2 (Homo sapiens)
Method: motif enrichment
not reviewed
Submitted by
Michael Snyder
Lab
Michael Snyder, Stanford
USF2 (Homo sapiens)
K562
Method: immunoprecipitation
Attachment from submitter
compliant
Caption
Arrow indicates immunoprecipitated band of expected size from K562 nuclear lysates. Lane 1 = K562 nuclear lysate. Lane 2 = K562 whole cell lysates. Lane 3 = supernatant from K562 immunoprecipitation (IP). Lane 4 = bound material from K562 IP. Lane 5 = bound material from control IgG IP from K562.
Submitted by
Kathrina Onate
Lab
Michael Snyder, Stanford
USF2 (Mus musculus)
Method: immunoprecipitation
not reviewed
Caption
Calculations were done by Pouya Kheradpour using a collection of known motifs (9 motifs were considered) available at http://www.broadinstitute.org/%7Epouyak/motif-disc/human/. Table 1 shows the fold-enrichments, enrichment p-values and fraction of peaks which contain the motif. The motif which produced the largest value for each criterion is shown in Table 1. Note that while the maximally enriched motif differs from the motif with the highest enrichment p-values and the most represented motif, the motifs are highly similar (Figure 1) and thus all values are similar between motifs. Motifs were identified using a matching stringency corresponding to 4-6 (6-mer). Peaks identified by IDR (1% cutoff) were used in the analysis and +/-50bp from peak centers were considered. Enrichments are for a given motif vs. a background consisting of +/- 50bp from the centers of all DNase I hypersensitive peaks. Repeat mask/simple repeats from UCSC and all gencode v7 exons (including non-protein coding genes) were excluded from the analysis. Comparison to shuffle motifs were used to correct for compositional bias. Enrichment is the corrected # of motifs in ChIP peaks/corrected # of motifs in DNaseI peaks. The current ENCODE standard calls for 4-fold enrichment and 10% motif representation for this criteria to be used for validation. All 5 USF2 datasets presented here easily exceed this threshold and the antibody is considered validated.
Submitted by
Michael Snyder
Lab
Michael Snyder, Stanford
USF2 (Mus musculus)
Method: motif enrichment
not reviewed
Submitted by
Michael Snyder
Lab
Michael Snyder, Stanford
USF2 (Homo sapiens)
K562GM12878HeLa-S3HepG2
Method: immunoblot
Attachment from submitter
compliant
Caption
Arrow indicates band consistent with expected size (37kD) of USF2 in whole cell lysates (from left to right): K562, GM12878, HeLa-S3, and HepG2 cell lines. A major band of ~45 kD is observed in all cell lines tested. This band is somewhat higher than expected for this protein (MW of 37 kD). However, this has been previously observed (see the image at http://www.abcam.com/USF2-antibody-ab60931.html) and may be due to post-translational modification. An additional band at ~100 kD is sometimes seen in the K562 and HepG2 cell lines, but this band is not immunoprecipitated efficiently (panel B).
Reviewer comment
In the K562 and HepG2 lanes, there is a more prominent band in the immunoblot near 100kD that subsequent IP-western in K562 shows, is not IP'd efficiently.
Submitted by
Kathrina Onate
Lab
Michael Snyder, Stanford