- MA3-046 - Invitrogen Antibodies KAT2A antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of GCN5 was performed by loading 10 µg of HeLa nuclear extract (lane 1), HeLa cytoplasmic extract (lane 2), U2OS nuclear extract (lane 3) and U2OS cytoplasmic extract (lane 4) in reducing sample buffer (Product # 39000) and Page Ruler Plus Protein Ladder (Product # 26619) onto a Novex 4-20% Tris-Glycine polyacrylamide gel (Product # WT4201BX10). Proteins were transferred to nitrocellulose membrane (Product # 88018) with Transfer Buffer (Product # 84731) using the G2 Gast Blotter (Product # 62288). Membrane was blocked in 5% Milk/TBST for one hour at room temperature. GCN5 was detected at approximately 94 kDa using GCN5 monoclonal antibody (Product # MA3-046) at a dilution of 1:2000 overnight at 4°C on a rocking platform, followed by a goat anti-mouse IgG-HRP secondary antibody (Product # 31430) at a dilution of 1:5,000 for one hour. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34076) and the myECL Imager (Product # 62236).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of GCN5 was performed by loading 10 µg of HeLa nuclear extract (lane 1), HeLa cytoplasmic extract (lane 2), U2OS nuclear extract (lane 3) and U2OS cytoplasmic extract (lane 4) in reducing sample buffer (Product # 39000) and Page Ruler Plus Protein Ladder (Product # 26619) onto a Novex 4-20% Tris-Glycine polyacrylamide gel (Product # WT4201BX10). Proteins were transferred to nitrocellulose membrane (Product # 88018) with Transfer Buffer (Product # 84731) using the G2 Gast Blotter (Product # 62288). Membrane was blocked in 5% Milk/TBST for one hour at room temperature. GCN5 was detected at approximately 94 kDa using GCN5 monoclonal antibody (Product # MA3-046) at a dilution of 1:2000 overnight at 4°C on a rocking platform, followed by a goat anti-mouse IgG-HRP secondary antibody (Product # 31430) at a dilution of 1:5,000 for one hour. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34076) and the myECL Imager (Product # 62236).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Chromatin immunoprecipitation analysis of HIV1 GCN5 was performed using cross-linked chromatin from Human 5A8 J-lat T lymphocytes culture latently infected with HIV1 and treated with 10 µg/mL PHA (phytohemaglutin) for 0, 4, and 8 hours. Immunoprecipitation was performed using a multiplex microplate Matrix ChIP assay (see reference for Matrix ChIP protocol: http://www.ncbi.nlm.nih.gov/pubmed/22098709) with 1.0 µL/100 µL well volume of GCN5 monoclonal antibody (Product # MA3-046). Chromatin aliquots from ~1 x 105 cells were used per ChIP pull-down. Quantitative PCR data were done in quadruplicate using 1 µL of eluted DNA in 2 µL SYBR real-time PCR reactions containing primers shown to amplify exon-1 of the Egr1 gene or HIV gag gene. PCR calibration curves were generated for each primer pair from a dilution series of sheared total genomic DNA. Quantitation of immunoprecipitated chromatin is presented as signal relative to the total amount of input chromatin. Results represent the mean +/- SEM for three experiments. Schematic representations of the Egr1 and HIV1 gag locus are shown above the data where boxes represent exons (black boxes = translated regions, white boxes = untranslated regions), the zigzag line represents an intron, and the straight line represents upstream sequence. Regions amplified by the primers are represented by black bars. Data courtesy of the Innovators Program.

MA3-046

Antibody data