重组Anti-KRAS抗体[EPR23474-76]

• IP

Unknown

Immunoprecipitation - Anti-KRAS antibody [EPR23474-76] (AB275876)

KRAS was immunoprecipitated from 0.35 mg Human brain tissue lysate with ab275876 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab275876 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Human brain tissue lysate 10 ug

Lane 2 : ab275876 IP in Human brain tissue lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab275876 in human brain tissue lysate

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 3 minutes.

All lanes:

Immunoprecipitation - Anti-KRAS antibody [EPR23474-76] (ab275876)

Predicted band size: 22 kDa

Observed band size: 21 kDa

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• WB

Lab

Western blot - Anti-KRAS antibody [EPR23474-76] (AB275876)

Anti-KRAS antibody [EPR23474-76] (ab275876) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab275876 was shown to bind specifically to KRAS. A band was observed at 21 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in KRAS knockout cell line ab276083 (knockout cell lysate ab284205). To generate this image, wild-type and KRAS knockout HCT 116 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5% BSA 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 VeriBlot for IP Detection Reagent (HRP) (ab131366) and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-KRAS antibody [EPR23474-76] (ab275876) at 1/1000 dilution

Lane 1:

Wild-type HCT 116 cell lysate at 10 µg

Lane 2:

KRAS knockout HCT 116 cell lysate at 10 µg

Observed band size: 21 kDa

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• WB

Lab

Western blot - Anti-KRAS antibody [EPR23474-76] (AB275876)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Exposure time : 3 minutes

All lanes:

Western blot - Anti-KRAS antibody [EPR23474-76] (ab275876) at 1/1000 dilution

Lane 1:

Human brain tissue lysate at 40 µg

Lane 2:

HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/50000 dilution

Predicted band size: 22 kDa

Observed band size: 21 kDa

false

• WB

Lab

Western blot - Anti-KRAS antibody [EPR23474-76] (AB275876)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Exposure time : 3 minutes.

All lanes:

Western blot - Anti-KRAS antibody [EPR23474-76] (ab275876) at 1/1000 dilution

Lane 1:

His-tagged human KRAS recombinant protein, 10 ng

Lane 2:

His-tagged human HRAS recombinant protein (aa 1-186), 20 ng

Lane 3:

His-tagged human NRAS recombinant protein (aa 1-186), 20 ng

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 22 kDa

Observed band size: 21 kDa

false