Anti-eIF4A2 抗体

• IP

Unknown

Immunoprecipitation - Anti-eIF4A2 antibody (AB31218)

eIF4A2 was immunoprecipitated using 0.5mg Mouse skeletal muscle whole tissue extract, 5μg of Rabbit polyclonal to eIF4A2 and 50μl of protein G magnetic beads (+). No antibody was added to the control (-).
The antibody was incubated under agitation with Protein G beads for 10min, Mouse skeletal muscle whole tissue extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
Proteins were eluted by addition of 40μl SDS loading buffer and incubated for 10min at 70^oC; 10μl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab31218.
Secondary : Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
Band : 47kDa : eIF4A2 .

All lanes:

Immunoprecipitation - Anti-eIF4A2 antibody (ab31218)

Predicted band size: 46 kDa

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• WB

Lab

Western blot - Anti-eIF4A2 antibody (AB31218)

Western blot : Anti-EIF4A2 antibody (ab31218) staining at 1 ug/ml, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab31218 was shown to bind specifically to EIF4A2. A band was observed at 40-50 kDa in wild-type A549 cell lysates with no signal observed at this size in EIF4A2 knockout cell line. To generate this image, wild-type and EIF4A2 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.

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Western blot - Anti-eIF4A2 antibody (ab31218) at 1 µg/mL

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

EIF4A2 knockout A549 cell lysate at 20 µg

Lane 3:

MCF7 cell lysate at 20 µg

Lane 4:

MCF7 Membrane Prep cell lysate at 20 µg

Secondary

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

Observed band size: 40 kDa,50 kDa

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• WB

Project

Western blot - Anti-eIF4A2 antibody (AB31218)

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Western blot - Anti-eIF4A2 antibody (ab31218) at 1 µg/mL

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Western blot - Jurkat whole cell lysate (<a href='/products/cell-lysates/jurkat-whole-cell-lysate-ab7899'>ab7899</a>) at 20 µg

Secondary

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IR Dye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/15000 dilution

Predicted band size: 46 kDa

Observed band size: 47 kDa,52 kDa

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• WB

Project

Western blot - Anti-eIF4A2 antibody (AB31218)

All lanes:

Western blot - Anti-eIF4A2 antibody (ab31218) at 1 µg/mL

Lane 1:

Brain (Mouse) Tissue Lysate at 10 µg

Lane 2:

Testis (Mouse) Tissue Lysate at 10 µg

Lane 3:

Western blot - Mouse skeletal muscle tissue lysate - total protein (<a href='/products/tissue-lysates/mouse-skeletal-muscle-tissue-lysate-total-protein-ab29711'>ab29711</a>) at 10 µg

Lane 4:

Spinal Cord (Mouse) Tissue Lysate at 10 µg

Lane 5:

Ovary (Mouse) Tissue Lysate at 10 µg

Lane 6:

PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate at 10 µg

Lane 7:

Brain (Rat) Tissue Lysate at 10 µg

Lane 8:

Heart (Rat) Tissue Lysate at 10 µg

Secondary

All lanes:

IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution

Predicted band size: 46 kDa

Observed band size: 47 kDa,51 kDa

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• WB

CiteAb

Western blot - Anti-eIF4A2 antibody (AB31218)

eIF4A2 western blot using anti-eIF4A2 antibody ab31218. Publication image and figure legend from Yan, L. X., Wu, Q. N., et al., 2011, Breast Cancer Res, PubMed 21219636.

ab31218 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab31218 please see the product overview.

ANKRD46 is a direct target of miR-21 in BC cell lines. (a) Predicted alignment of miR-21 with the target site derived from ANKRD46 and EIF4A2 3' UTR, determined with the software miRBase Targets V5 and miRNAMap, respectively. Note the seed matches at the 5' end of miR-21 (grey boxes) and the mutated nucleotides (underlined). (b) Luciferase assays show that miR-21 directly repress ANKRD46 mRNAs through 3' UTR interactions. Part of the 3' UTRs of wild-type ANKRD46 (478-bp length) and EIF4A2 (194-bp length), or the mutations were cloned into pMIR-REPORT vector (Applied Biosystems), downstream of luciferase. These vectors were then cotransfected with synthetic miR-21 (pre-miR-21) or miR-control in 293T cells, and luciferase activity was quantified. The graph shows the percentage of remaining luciferase activity calculated by normalizing the miR-21 expression values on the miR-control values. (c) Western blot to assay ANKRD46 and EIF4A2 after miR-21 knockdown, with GAPDH as equal loading control, followed by densitometric analysis. Cells were treated and harvested as described in Figure 4b. Data in (b) and (c) are mean + SD (n = 3). * p < 0.05. WT, wild-type; Mut, mutant.

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