Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] - BSA and Azide free

• WB

Supplier Data

Western blot - Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] - BSA and Azide free (AB315091)

This data was developed using ab315090, the same antibody clone in a different buffer formulation.

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

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

Exposure time : Lane 1 : 81 seconds; Lane2-3 : 180 seconds

All lanes:

Western blot - Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] (<a href='/en-us/products/primary-antibodies/pip5k3-pikfyve-antibody-epr27944-21-ab315090'>ab315090</a>) at 1/1000 dilution

Lane 1:

THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Lane 2:

RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 3:

NR8383 (rat lung macrophage (alveolar)) whole cell lysate at 20 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

Observed band size: 250 kDa

false

• WB

Supplier Data

Western blot - Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] - BSA and Azide free (AB315091)

This data was developed using ab315090, the same antibody clone in a different buffer formulation.

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

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] (<a href='/en-us/products/primary-antibodies/pip5k3-pikfyve-antibody-epr27944-21-ab315090'>ab315090</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

HeLa transfected with siRNA specifically targeting PIKFYVE whole cell lysate at 20 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

Observed band size: 250 kDa,124 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] - BSA and Azide free (AB315091)

This data was developed using ab315090, the same antibody clone in a different buffer formulation.

Western blot : Anti-PIKFYVE antibody (ab315090) 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, ab315090 was shown to bind specifically to PIKFYVE. A band was observed at 237 kDa in wild-type A549 cell lysates with no signal observed at this size in PIKFYVE knockout cell line. To generate this image, wild-type and PIKFYVE 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-PIP5K3/PIKFYVE antibody [EPR27944-21] (<a href='/en-us/products/primary-antibodies/pip5k3-pikfyve-antibody-epr27944-21-ab315090'>ab315090</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

Western blot - Human PIKFYVE Knockout A549 cell line (ab314965)

Lane 2:

PIKFYVE knockout A549 cell lysate at 20 µg

Lane 3:

HeLa UV treated (100 mJ/cm^2, 1 hr recovery, new, Branford) cell lysate at 20 µg

Lane 4:

HeLa UV treated (40 mJ/cm^2 30 min recovery), LAMBDA PHOS 4X, 2h cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Goat anti-Rabbit IgG H&L 800CW at 1/20000 dilution

Lanes 1 - 4:

Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Observed band size: 237 kDa

false

• WB

Supplier Data

Western blot - Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] - BSA and Azide free (AB315091)

This data was developed using ab315090, the same antibody clone in a different buffer formulation.

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

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-PIP5K3/PIKFYVE antibody [EPR27944-21] (<a href='/en-us/products/primary-antibodies/pip5k3-pikfyve-antibody-epr27944-21-ab315090'>ab315090</a>) at 1/1000 dilution

Lane 1:

Mouse liver tissue lysate

Lane 2:

Mouse testis tissue lysate

Lane 3:

Mouse spleen tissue lysate

Lane 4:

Rat testis tissue lysate

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

Observed band size: 250 kDa,124 kDa

false

Exposure time: 180s