Anti-KAT2A / GCN5 antibody [EPR21146] - BSA and Azide free

• IP

Supplier Data

Immunoprecipitation - Anti-KAT2A / GCN5 antibody [EPR21146] - BSA and Azide free (AB231075)

KAT2A / GCN5 was immunoprecipitated from 0.35mg of HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab217876 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab217876 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

Lane 1 : HeLa whole cell lysate 10ug (Input).

Lane 2 : ab217876 IP in HeLa whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab217876 in HeLa whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 30 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217876).

All lanes:

Immunoprecipitation - Anti-KAT2A / GCN5 antibody [EPR21146] - ChIP Grade (<a href='/en-us/products/primary-antibodies/kat2a-gcn5-antibody-epr21146-chip-grade-ab217876'>ab217876</a>)

Predicted band size: 94 kDa

Observed band size: 94 kDa

false

• ChIP

Unknown

ChIP - Anti-KAT2A / GCN5 antibody [EPR21146] - BSA and Azide free (AB231075)

Chromatin was prepared from A549 treated with thapsigargin (1μM 12 hours) cells according to the Abcam Dual X-ChIP protocol*. Cells were fixed with EGS for 30 minutes, then formaldehyde for 10 minutes.
The ChIP was performed with 25 μg of chromatin, 5 μg of ab217876 (red), and 20 μl of Protein A/G sepharose beads. 5 μg of rabbit normal IgG was added to the beads control (gray). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

Primers and probes are located in the first kb of the transcribed region.
*http : //www.abcam.com/resources?keywords=X%20ChIP%20protocol

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217876).

• WB

Supplier Data

Western blot - Anti-KAT2A / GCN5 antibody [EPR21146] - BSA and Azide free (AB231075)

Blocking/Dilution buffer : 5% NFDM/TBST

Exposure time : 3 minutes

ab217876 was shown to specifically react with KAT2A in wild type HAP1 cells. No band was observed when KAT2A knockout samples were used. Wild-type and KAT2A knockout samples were subjected to SDS-PAGE. ab217876 and ab181602 (Human anti-GAPDH loading control) were incubated overnight at 4°C at a 1/1000 dilution and 1/5000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100,000 dilution for 1 hour at room temperature before imaging. The blot was developed on a BIO-RAD® ChemiDoc™ MP instrument.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217876).

All lanes:

Western blot - Anti-KAT2A / GCN5 antibody [EPR21146] - BSA and Azide free (ab231075) at 1/1000 dilution

Lane 1:

Wild type HAP1 whole cell lysate at 10 µg

Lane 2:

KAT2A / GCN5-knockout HAP1 whole cell lysate whole cell lysate at 10 µg

Lane 3:

HEK-293T (uman epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 10 µg

Lane 4:

MCF7 (human breast adenocarcinoma cell line) whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 94 kDa

true

• WB

Supplier Data

Western blot - Anti-KAT2A / GCN5 antibody [EPR21146] - BSA and Azide free (AB231075)

False colour image of Western blot : Anti-KAT2A / GCN5 antibody [EPR21146] - ChIP Grade staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab217876 was shown to bind specifically to KAT2A / GCN5. A band was observed at 95 kDa in wild-type U-2 OS cell lysates with no signal observed at this size in kat2a knockout cell line. To generate this image, wild-type and kat2a knockout U-2 OS cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution. This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217876).

All lanes:

Western blot - Anti-KAT2A / GCN5 antibody [EPR21146] - ChIP Grade (<a href='/en-us/products/primary-antibodies/kat2a-gcn5-antibody-epr21146-chip-grade-ab217876'>ab217876</a>) at 1/1000 dilution

Lane 1:

Wild-type U-2 OS cell lysate at 20 µg

Lane 2:

GCN5 knockout U-2 OS cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

NIH/3T3 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 94 kDa

Observed band size: 95 kDa

false