ENCAB426WVA

Antibody against Homo sapiens ZMYM3

Homo sapiens
HepG2, K562, MCF-7, GM12878
characterized to standards with exemption
Homo sapiens
any cell type or tissue
partially characterized
Status
released
Source (vendor)
CDI
Product ID
JH39.2.2F10
Lot ID
HAIB-001
Characterized targets
ZMYM3 (Homo sapiens)
Host
mouse
Clonality
monoclonal
Purification
Protein G
Isotype
IgG2a
Antigen description
Monospecific for human ZMYM3
External resources

Characterizations

ZMYM3 (Homo sapiens)
Method: immunoprecipitation followed by mass spectrometry
Attachment from submitter
not compliant
Caption
K562 whole cell lysate was immunoprecipitated using the primary antibody (CDI; JH39.2.2F10). The IP fraction was loaded on a 12% Bio-Rad TGX gel and separated with the Bio-Rad Tetra Cell system. Gel fragments (rectangle outline) corresponding to the bands indicated on the Coomassie Blue stained gel image were excised and sent to the University of Alabama at Birmingham Cancer Center Mass Spectrometry/Proteomics Shared Facility. Analysis of gel fragment 1 from K562: The sample was analyzed on a LTQ XL Linear Ion Trap Mass Spectrometer by LC-ESI-MS/MS. Peptides were identified using SEQUEST tandem mass spectral analysis with probability based matching at p < 0.05. SEQUEST results were reported with ProteinProphet protXML Viewer (TPP v4.4 JETSTREAM) and filtered for a minimum probability of 0.9. All protein hits that met these criteria were reported, including common contaminants. Fold enrichment for each protein reported was determined using a custom script based on the FC-B score calculation from the reference Mellacheruvu et al., 2013. The CRAPome: a contaminant repository for affinity purification mass spectrometry data. Nat. Methods. 10(8):730-736. Doi:10.1038/nmeth.2557. The target protein, ZMYM3, was identified as the 26th enriched protein and the 2nd transcription factor based on IP-Mass Spectrometry.
Reviewer comment
Factor of interest is not within the top 20 proteins ranked by fold enrichment
Submitted by
Mark Mackiewicz
Lab
Richard Myers, HAIB
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ZMYM3 (Homo sapiens)
Method: immunoprecipitation followed by mass spectrometry
Attachment from submitter
compliant
Caption
K562 whole cell lysate was immunoprecipitated using the primary antibody (CDI; JH39.2.2F10). The IP fraction was loaded on a 12% Bio-Rad TGX gel and separated with the Bio-Rad Tetra Cell system. Gel fragments (rectangle outline) corresponding to the bands indicated on the Coomassie Blue stained gel image were excised and sent to the University of Alabama at Birmingham Cancer Center Mass Spectrometry/Proteomics Shared Facility. Analysis of gel fragment 2 from K562: The sample was analyzed on a LTQ XL Linear Ion Trap Mass Spectrometer by LC-ESI-MS/MS. Peptides were identified using SEQUEST tandem mass spectral analysis with probability based matching at p < 0.05. SEQUEST results were reported with ProteinProphet protXML Viewer (TPP v4.4 JETSTREAM) and filtered for a minimum probability of 0.9. All protein hits that met these criteria were reported, including common contaminants. Fold enrichment for each protein reported was determined using a custom script based on the FC-B score calculation from the reference Mellacheruvu et al., 2013. The CRAPome: a contaminant repository for affinity purification mass spectrometry data. Nat. Methods. 10(8):730-736. Doi:10.1038/nmeth.2557. The target protein, ZMYM3, was identified as the 1st enriched protein and the 1st transcription factor based on IP-Mass Spectrometry.
Submitted by
Mark Mackiewicz
Lab
Richard Myers, HAIB
Download
ZMYM3 (Homo sapiens)
Method: immunoprecipitation
Attachment from submitter
Reviewer comment
IP gel submitted for mass spectrometry analysis
Submitted by
Mark Mackiewicz
Lab
Richard Myers, HAIB
ZMYM3 (Homo sapiens)
K562GM12878HepG2MCF-7
Method: immunoprecipitation
Attachment from submitter
exempt from standards
Caption
Whole cell lysates of K562, MCF7, HepG2 and GM12878 were immunoprecipitated using the primary antibody (CDI; JH39.2.2F10). The IP fraction was separated on a 12% acrylamide gel with the Bio-Rad PROTEAN II xi system. After separation, the samples were transferred to a nitrocellulose membrane with an Invitrogen iBlot system. The membrane was probed with the primary antibody (same as that used for IP) and a secondary HRP-conjugated antibody. The resulting bands were visualized with SuperSignal West Femto Solution (Thermo Scientific). Protein Marker (PM) is labeled in kDa. Two bands were detected at ~150 and 220 kDa. Expected size ~152 kDa.
Submitter comment
--
Reviewer comment
Rescued by mass spectrometry analysis
Submitted by
Mark Mackiewicz
Lab
Richard Myers, HAIB
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