ENCAB000BNW

Antibody against Homo sapiens SKIL

Homo sapiens
K562, GM12878
characterized to standards
Homo sapiens
any cell type or tissue
partially characterized
Homo sapiens
liver, HepG2
not characterized to standards
Status
released
Source (vendor)
Sigma
Product ID
HPA008472
Lot ID
R02251
Characterized targets
SKIL (Homo sapiens)
Host
rabbit
Clonality
polyclonal
Purification
affinity
Isotype
IgG
Antigen description
Ski-like protein recombinant protein epitope signature tag (PrEST)
Antigen sequence
SPDKRTCHWGFESAKWHCYLHVNQKYLGTPEEKKLKIILEEMKEKFSMRSGKRNQSKTDAPSGMELQSWYPVIKQEGDHVSQTHSFLHPSYYLYMCDKVVAPNVSLTSAVSQSKELTKTEASKSISRQSEKAHSSGKLQ
External resources

Characterizations

SKIL (Homo sapiens)
Method: immunoprecipitation
Attachment from submitter
Caption
Immunoprecipitation was performed on nuclear extracts from the cell line: MCF-7, using the antibody HPA008472. The blot shows western blot analysis of input, flowthrough, immunoprecipitate and mock immunoprecipitate using IgG.Molecular Weight: 77.0
Submitted by
Denis Salins
Lab
Michael Snyder, Stanford
SKIL (Homo sapiens)
Method: immunoprecipitation
Attachment from submitter
Caption
Immunoprecipitation was performed on nuclear extracts from the cell line: HepG2, using the antibody HPA008472. The blot shows western blot analysis of input, flowthrough, immunoprecipitate and mock immunoprecipitate using IgG.Molecular Weight: 77.0
Submitted by
Denis Salins
Lab
Michael Snyder, Stanford
SKIL (Homo sapiens)
Method: immunoprecipitation
Attachment from submitter
Caption
Immunoprecipitation was performed on nuclear extracts from the cell line: GM12878, using the antibody HPA008472. The blot shows western blot analysis of input, flowthrough, immunoprecipitate and mock immunoprecipitate using IgG.Molecular Weight: 77.0
Submitted by
Denis Salins
Lab
Michael Snyder, Stanford
SKIL (Homo sapiens)
Method: immunoprecipitation
Attachment from submitter
Caption
Immunoprecipitation was performed on nuclear extracts from the cell line: HEK293T, using the antibody HPA008472. The blot shows western blot analysis of input, flowthrough, immunoprecipitate and mock immunoprecipitate using IgG.Molecular Weight: 77.0
Submitted by
Nathaniel Watson
Lab
Michael Snyder, Stanford
SKIL (Homo sapiens)
K562
Method: immunoprecipitation
Attachment from submitter
exempt from standards
Caption
Immunoprecipitation was performed on nuclear extracts from the cell line K562 using the antibody HPA008472. Lane 1: input nuclear lysate. Lane 2: material immunoprecipitated with antibody. Lane 3: material immunoprecipitated using control IgG. Marked bands were excised from gel and subjected to analysis by mass spectrometry. Target molecular weight: 77.0.
Submitter comment
We analyzed the whole marked band, including what looks to be immunoreactive signal in the IgG control.
Reviewer comment
Marked band looks to also be detected in the IgG control. However, in the subsequent mass-spec analysis, SKIL is found to be specifically enriched in the IP and not in the IgG control.
Submitted by
Nathaniel Watson
Lab
Michael Snyder, Stanford
SKIL (Homo sapiens)
Method: immunoprecipitation followed by mass spectrometry
compliant
Caption
IP followed by mass spectrometry. Briefly, protein was immunoprecipitated from K562 nuclear cell lysates using the antibody HPA008472, and the IP fraction was loaded on a 10% polyacrylamide gel (NuPAGEBis-Tris Gel) and separated with an Invitrogen NuPAGE electrophoresis system. The gel was stained by ColloidialCoomassie G-250 stain, gel fragments corresponding to the bands indicated were excised. Then proteins were trypsinized using the in-gel digestion method. Digested proteins were analyzed on an Orbitrap Elite mass spectrometer (Thermo Scientific) by the nanoLC-ESI-MS/MS technique. Peptides were identified by the SEQUEST algorithm and filtered with a high confidence threshold (Peptide false discovery rate < 1%, 2 unique peptides per protein minimum, mass error < 10 ppm).
Submitter comment
SHMT2, KIF20A, MORC3, SF3B3, PARN, SUGP1, CIZ1 and G3BP1 are not DNA binding transcription factors.
Reviewer comment
CIZ1 might be a sequence-specific TF? https://www.ncbi.nlm.nih.gov/pubmed/12824700. However, the ENCODE Antibody Review panel reviewed the reference on June 26, 2017 and determined that more recent papers consider CIZ1 a co-activator and the degenerate consensus motif cited in the reference is not convincing evidence of it being a sequence-specific DNA binding protein.
Submitted by
Nathaniel Watson
Lab
Michael Snyder, Stanford
SKIL (Homo sapiens)
GM12878K562HepG2liver
Method: immunoblot
Attachment from submitter
compliant
Caption
a) Western blot analysis of nuclear lysates prepared from multiple cells lines loaded in the order: GM12878, K562, HepG2, Liver using the antibody HPA008472.
Reviewer comment
Major band smaller than expected size of ~77kD (not indicated on image or caption) but there are multiple bands.
Submitted by
Trupti Kawli
Lab
Michael Snyder, Stanford
SKIL (Homo sapiens)
K562
Method: immunoprecipitation
Attachment from submitter
compliant
Caption
b) Immunoprecipitation was performed on nuclear extracts from the cell line: K562, using the antibody HPA008472 .The blot shows western blot analysis of input, flowthrough, immunoprecipitate and mock immunoprecipitate using IgG. Expected size ~77 kDa.
Submitted by
Trupti Kawli
Lab
Michael Snyder, Stanford