Anti-PERK antibody

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-PERK antibody (AB65142)

Immunofluorescence analysis of HepG2 cells using ab65142 at a 1/500-1/1000 dilution. Left image untreated. Right image treated with immunising peptide.

• WB

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Western blot - Anti-PERK antibody (AB65142)

The amount of positive control loading for the WB is 5-30 ug of total protein.
The amount of the peptide for the WB is 5-10 ug.

All lanes:

Western blot - Anti-PERK antibody (ab65142) at 1/500 dilution

Lane 1:

Extracts from Hela cells

Lane 2:

Extracts from HepG2 cells

Lane 3:

Extracts from HepG2 cells, plus immunising peptide

Predicted band size: 125 kDa

Observed band size: 125 kDa

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• WB

Lab

Western blot - Anti-PERK antibody (AB65142)

Western blot : Anti-EIF2AK3 antibody (ab65142) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab65142 was shown to bind specifically to EIF2AK3. A band was observed at 145 kDa in wild-type A549 cell lysates with no signal observed at this size in EIF2AK3 knockout cell line. To generate this image, wild-type and EIF2AK3 knockout A549 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.

All lanes:

Western blot - Anti-PERK antibody (ab65142) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

EIF2AK3 knockout A549 cell lysate at 20 µg

Lane 2:

Western blot - Human EIF2AK3 knockout A549 cell line (<a href='/en-us/products/cell-lines/human-eif2ak3-knockout-a549-cell-line-ab288905'>ab288905</a>)

Lane 3:

MCF7 cell lysate at 20 µg

Lane 4:

Human eye 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

Observed band size: 145 kDa

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