1. Antibodies
  2. Homo sapiens
  3. SREBF2

ENCAB000ALD

Antibody against Homo sapiens SREBF2

Homo sapiens
K562, HeLa-S3, HepG2, IMR-90, A549
characterized to standards
Homo sapiens
any cell type or tissue
partially characterized
Homo sapiens
GM12878, heart
not characterized to standards
Status
released
Source (vendor)
Cayman
Product ID
10007663
Lot ID
0424883-1
Characterized targets
SREBF2 (Homo sapiens)
Host
rabbit
Clonality
polyclonal
Purification
affinity

Characterizations

SREBF2 (Homo sapiens)
Method: immunoprecipitation followed by mass spectrometry
Attachment PDF Icon
not reviewed
Caption
Immunoprecipitation of SREBP2 from HepG2 cells using 10007663 antibody. Lane 1: input nuclear lysate, Lane 2: material immunoprecipitated with 10007663, Lane 3: material immunoprecipitated using control IgG. Bands A was excised from the gel and subjected to analysis by mass spectrometry. IP followed by masspectrometry: Briefly, protein was immunoprecipitated from HepG2 whole cell lysates using 10007663, and the IP fraction was loaded on a 10% polyacrylamide gel (NuPAGE Bis-Tris Gel) and separated with an Invitrogen NuPAGE electrophoresis system. The gel was silver-stained, gel fragments corresponding to the bands indicated were excised and destained using the SilverSNAP Stain for Mass Spectrometry (Pierce). Then proteins were trypsinized using the in-gel digestion method. Digested proteins were analyzed on an LTQ- Orbitrap (Thermo Scientific) by the nanoLC-ESI-MS/MS technique. Peptides were identified by the SEQUEST algorithm and filtered with a high confidence threshold (Protein false discovery rate < 1%, 2 peptides per protein minimum)."! We report 26 proteins identified in band A, although 3 of these are also present in a control immunoprecipitation and are thus likely to present due to non-specific association with the IP matrix. Of the specifically immunoprecipitated proteins, SREBP2 is the most abundant (185 peptides).! Based on these observations, this band is likely due to the presence of immunoprecipitated SREBP2 and 10007663 meets the ENCODE standard for validation by this criterion.
Submitted by
Michael Snyder
Lab
Michael Snyder, Stanford
SREBF2 (Homo sapiens)
Method: immunoblot
Attachment PDF Icon
not reviewed
Caption
Western blots on nuclear lysates from cell lines GM12878, K562, HeLaS3, HepG2, IMR90, human heart tissue and A549. The antibody detects proteins ~60 kD and higher than 125kD as indicated in manufacturer product sheet. Bands in the expected size range were observed in multiple human cell types. The bands detected by anti SREBP2 antibody (10007663) were analyzed further by IP-Mass Spec.
Submitted by
Michael Snyder
Lab
Michael Snyder, Stanford
SREBF2 (Homo sapiens)
HepG2
Method: immunoprecipitation
Attachment from submitter
Attachment from submitters
compliant
Caption
Immunoprecipitation of SREBP2 from HepG2 cells using 10007663 antibody. Lane 1: input nuclear lysate, Lane 2: material immunoprecipitated with 10007663, Lane 3: material immunoprecipitated using control IgG. Bands A was excised from the gel and subjected to analysis by mass spectrometry.
Reviewer comment
How come both the smaller active form (50-68kD) and the larger (~126 kD) were excised in the same band (A) for mass spec?
Submitted by
Kathrina Onate
Lab
Michael Snyder, Stanford
SREBF2 (Homo sapiens)
Method: immunoprecipitation followed by mass spectrometry
Attachment from submitter
Attachment from submitters
compliant
Caption
IP followed by mass spectrometry: Briefly, protein was immunoprecipitated from HepG2 whole cell lysates using 10007663, and the IP fraction was loaded on a 10% polyacrylamide gel (NuPAGE Bis-Tris Gel) and separated with an Invitrogen NuPAGE electrophoresis system. The gel was silver-stained, gel fragments corresponding to the bands indicated were excised and destained using the SilverSNAP Stain for Mass Spectrometry (Pierce). Then proteins were trypsinized using the in-gel digestion method. Digested proteins were analyzed on an LTQ- Orbitrap (Thermo Scientific) by the nanoLC-ESI-MS/MS technique. Peptides were identified by the SEQUEST algorithm and filtered with a high confidence threshold (Protein false discovery rate < 1%, 2 peptides per protein minimum)."! We report 26 proteins identified in band A, although 3 of these are also present in a control immunoprecipitation and are thus likely to present due to non-specific association with the IP matrix. Of the specifically immunoprecipitated proteins, SREBP2 is the most abundant (185 peptides).
Submitted by
Kathrina Onate
Lab
Michael Snyder, Stanford
SREBF2 (Homo sapiens)
GM12878K562HeLa-S3HepG2IMR-90heartA549
Method: immunoblot
Attachment from submitter
Attachment from submitters
compliant
Caption
Western blots on nuclear lysates from cell lines GM12878, K562, HeLaS3, HepG2, IMR90, human heart tissue and A549. The antibody detects proteins ~60 kD and higher than 125kD as indicated in manufacturer product sheet. Bands in the expected size range were observed in multiple human cell types. The bands detected by anti SREBP2 antibody (10007663) were analyzed further by IP-Mass Spec.
Reviewer comment
Expected size of ~126kD sometimes runs larger because of glycosylation. Active form is cleaved and is ~50-68kD: https://www.caymanchem.com/app/template/Product.vm/catalog/10007663
Submitted by
Kathrina Onate
Lab
Michael Snyder, Stanford