1. Antibodies
  2. Homo sapiens
  3. NANOG

ENCAB000AIX

Antibody against Homo sapiens NANOG

Homo sapiens
H1, GM23338
characterized to standards with exemption
Status
released
Source (vendor)
Santa Cruz Biotech
Product ID
sc-33759
Lot ID
C0909
Characterized targets
NANOG (Homo sapiens)
Host
rabbit
Clonality
polyclonal
Isotype
IgG
Antigen description
Epitope corresponding to amino acids 151-305 mapping at the C-terminus of Nanog of human origin.

Characterizations

NANOG (Homo sapiens)
Method: immunoprecipitation followed by mass spectrometry
Attachment PDF Icon
exempt from standards
Caption
IP followed by mass spectrometry: Briefly, human ES whole cell lysates were immunoprecipitated using primary antibody, and the IP fraction was loaded on a 12% acrylamide gel and separated with a Bio-Rad PROTEAN II xi system. Gel was stained with Coomassie Blue in order to visualize marker bands. A gel fragment corresponding to the band indicated above in the western blot image was excised and sent to the University of Alabama at Birmingham Cancer Center Mass Spectrometry/Proteomics Shared Facility. There the sample was run on an LTQ XL Linear Ion Trap Mass Spectrometer by LC-ESI-MS/MS. Peptides were identified using SEQUEST tandem mass spectra analysis, with probability based matching at p < 0.05. As per ENCODE data standards, all SEQUEST results are attached, including common contaminants. Target protein is listed as hit 5, highlighted in bold font.
Submitter comment
This antibody was characterizes to ENCODE2 standards.
Reviewer comment
I think that the IP and mass spec are good enough to be classified as an exemption characterized to ENCODE2 standards.
Submitted by
Richard Myers
Lab
Richard Myers, HAIB
NANOG (Homo sapiens)
H1
Method: immunoblot
Attachment from submitter
Attachment from submitters
exempt from standards
Caption
Western blot protocol: Whole cell lysate was immunoprecipitated using primary antibody, and the IP fraction was loaded on a 12% acrylamide gel and separated with a Bio-Rad PROTEAN II xi system. After separation, the samples were transferred to a nitrocellulose membrane with an Invitrogen iBlot system. Blotting with primary (same as that used for IP) and secondary HRP-conjugated antibodies was performed on an Invitrogen BenchPro 4100 system. Visualization was achieved using SuperSignal West Femto solution (Thermo Scientific). Results: Band of expected size visualized, representing strongest signal in the lane. Figure legend: IP-western with sc-33759 in whole cell lysates of human ES cells. NANOG band is indicated, as is heavy chain of IgG.
Submitter comment
This antibody was characterizes to ENCODE2 standards.
Reviewer comment
DCC: Band of interest is not within 20% of the expected size PJF:I think that the IP and mass spec are good enough to be classified as an exemption characterized to ENCODE2 standards.
Submitted by
Richard Myers
Lab
Richard Myers, HAIB
NANOG (Homo sapiens)
GM23338
Method: immunoblot
Attachment from submitter
Attachment from submitters
exempt from standards
Caption
The ENCODE Binding Working Group finds for some valuable biosamples that recreating a primary on well characterized antibodies is not cost effective. Therefore, they allow exemption from standards for these samples.
Submitter comment
The lab is asking for an exemption forIPSCs due to the lack of resources to make a primary characterization for them
Reviewer comment
Exempted by the Feb 29, 2016 antibody review panel. Standards require characterizations exempt due to biosample shortage remain pending until they get compliant primary characterizations from two different cell types.
Submitted by
Richard Myers
Lab
Richard Myers, HAIB