Anti-Axl antibody [EPR21107]

• WB

Supplier Data

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

Exposure time : Lane 1 : 6 seconds; Lane 2 : 3 seconds; Lane 3 : 46 seconds; Lanes 4-5 : 3 minutes; Lanes 6-7 : 10 seconds.

Blocking/Dilution buffer : 5% NFDM/TBST.

The 140 kDa band corresponds to the full-length Axl, while the 80 kDa band could be the cleaved form of Axl (PMID : 7822279 and 19541935).

The apparent molecular mass is higher than predicted, likely due to glycosylation (PMID : 23629654).

All lanes:

Western blot - Anti-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lane 1:

Human skeletal muscle lysate at 20 µg

Lane 2:

Rat muscle lysate at 10 µg

Lane 3:

Mouse muscle lysate at 10 µg

Lane 4:

4T1 (mouse mammary gland carcinoma cell line) whole cell lysate at 20 µg

Lane 5:

C2C12 (mouse myoblast cell line) whole cell lysate at 20 µg

Lane 6:

HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 7:

HUVEC (human umbilical vein endothelial cell line) 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 1/50000 dilution

Predicted band size: 98 kDa

Observed band size: 140 kDa,80 kDa

true

• WB

Lab

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

Lanes 1 - 4 : Merged signal (red and green). Green - ab215205 observed at 135 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55kDa.

ab215205 was shown to react with Axl in western blot. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab215205 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4°C at a 1 in 1000 Dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 40 µg

Lane 2:

AXL knockout HeLa cell lysate at 40 µg

Lane 3:

Wild-type HeLa cell lysate at 20 µg

Lane 4:

AXL knockout HeLa cell lysate at 20 µg

Predicted band size: 98 kDa

Observed band size: 135 kDa

false

• WB

Unknown

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

Western blot : Anti-Axl antibody [EPR21107] (ab215205) staining at 1 : 1000 dilution, shown in green; Anti-GAPDH antibody [6C5] - Loading Control (ab8245) shown in red.

ab215205 was shown to bind specifically to Axl. A band was observed at 140 kDa in wild-type HeLa cell lysates with no signal observed at this size in Axl CRISPR-Cas9 edited cell line ab261810 (CRISPR-Cas9 edited cell lysate ab257151). The band observed in the CRISPR-Cas9 edited lysate lane below 90 kDa is likely to represent a truncated form of Axl. This has not been investigated further and the functional properties of the gene product have not been determined.

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 (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/10000 dilution.

All lanes:

Western blot - Anti-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human AXL knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-axl-knockout-hela-cell-line-ab261810'>ab261810</a>)

Lane 2:

Western blot - Human AXL knockout HeLa cell lysate (<a href='/en-us/products/cell-lysates/human-axl-knockout-hela-cell-lysate-ab257151'>ab257151</a>)

Lane 2:

AXL knockout HeLa cell lysate at 20 µg

Lane 3:

NCI-H1299 cell lysate at 20 µg

Lane 4:

Jurkat cell lysate at 20 µg

Predicted band size: 98 kDa

Observed band size: 80140 kDa

false

• WB

Supplier Data

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

Blocking/Dilution buffer : 5% NFDM/TBST.

Negative control : JurkatThe 140 kDa band corresponds to the full-length Axl, while the 80 kDa band could be the cleaved form of Axl (PMID : 7822279 and 19541935).

The apparent molecular mass is higher than predicted, likely due to glycosylation (PMID : 23629654).

All lanes:

Western blot - Anti-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lane 1:

DU 145 (human prostate carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

NCI-H1299 (human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 3:

Jurkat (human T cell leukemia cell line from peripheral blood) 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 1/50000 dilution

Predicted band size: 98 kDa

Observed band size: 140 kDa,80 kDa

true

Exposure time: 29s

• WB

Lab

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

Western blot : Anti-Axl antibody [EPR21107] (ab215205) staining at 1 : 1000 dilution, shown in green; Anti-GAPDH antibody [6C5] - Loading Control (ab8245) shown in red.

ab215205 was shown to bind specifically to Axl. A band was observed at 140 kDa in wild-type HeLa cell lysates with no signal observed at this size in Axl CRISPR-Cas9 edited cell line ab265392 (CRISPR-Cas9 edited cell lysate ab257152). The band observed in the CRISPR-Cas9 edited lysate lane below 90 kDa is likely to represent a truncated form of Axl. This has not been investigated further and the functional properties of the gene product have not been determined.

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 (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/10000 dilution.

All lanes:

Western blot - Anti-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

AXL knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human AXL knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-axl-knockout-hela-cell-line-ab265392'>ab265392</a>)

Lane 2:

Western blot - Human AXL knockout HeLa cell lysate (<a href='/en-us/products/cell-lysates/human-axl-knockout-hela-cell-lysate-ab257152'>ab257152</a>)

Lane 3:

NCI-H1299 cell lysate at 20 µg

Lane 4:

Jurkat cell lysate at 20 µg

Predicted band size: 98 kDa

Observed band size: 80140 kDa

false

• WB

Lab

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

Western blot : Anti-Axl antibody [EPR21107] (ab215205) staining at 1 : 1000 dilution, shown in green; Anti-GAPDH antibody [6C5] - Loading Control (ab8245) shown in red.

ab215205 was shown to bind specifically to Axl. A band was observed at 140 kDa in wild-type HeLa cell lysates with no signal observed at this size in Axl CRISPR-Cas9 edited cell line ab261810 (CRISPR-Cas9 edited cell lysate ab257151). The band observed in the CRISPR-Cas9 edited lysate lane below 90 kDa is likely to represent a truncated form of Axl. This has not been investigated further and the functional properties of the gene product have not been determined.

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 (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/10000 dilution.

All lanes:

Western blot - Anti-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

AXL knockout HeLa cell lysate at 20 µg

Lane 3:

NCI-H1299 cell lysate at 20 µg

Lane 4:

Jurkat cell lysate at 20 µg

Predicted band size: 98 kDa

Observed band size: 140 kDa

false

• WB

Lab

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

False colour image of Western blot : Anti-Axl antibody [EPR21107] 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, ab215205 was shown to bind specifically to Axl. A band was observed at 135 kDa in wild-type A549 cell lysates with no signal observed at this size in AXL knockout cell line ab273744 (knockout cell lysate ab273780). To generate this image, wild-type and AXL 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-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

AXL knockout A549 cell lysate at 20 µg

Lane 2:

Western blot - Human AXL knockout A549 cell line (<a href='/en-us/products/cell-lines/human-axl-knockout-a549-cell-line-ab273744'>ab273744</a>)

Lane 3:

A549 cell lysate at 20 µg

Lane 4:

Jurkat cell lysate at 20 µg

Predicted band size: 98 kDa

Observed band size: 135 kDa

false

• WB

Lab

Western blot - Anti-Axl antibody [EPR21107] (AB215205)

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

ab219651 exhibits low sensitivity in western blot. It is recommended to use ab259831 or ab215205 as alternatives or to optimize experimental conditions by increasing the sample loading amount, using a lower antibody dilution ratio, and employing femtogram-level sensitivity substrates.

Lanes 1 - 3:

Western blot - Anti-Axl antibody [EPR19880] (<a href='/en-us/products/primary-antibodies/axl-antibody-epr19880-ab219651'>ab219651</a>) at 1/1000 dilution

Lanes 5 - 7:

Western blot - Anti-Axl antibody [EPR23892-15] (<a href='/en-us/products/primary-antibodies/axl-antibody-epr23892-15-ab259831'>ab259831</a>) at 1/1000 dilution

Lanes 9 - 11:

Western blot - Anti-Axl antibody [EPR21107] (ab215205) at 1/1000 dilution

Lanes 1, 5 and 9:

HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 20 µg

Lanes 2, 6 and 9:

HUVEC (human umbilical vein endothelial cell line) whole cell lysate 20 µg

Lanes 3, 7 and 11:

A549 (Human lung carcinoma epithelial cell) whole cell lysate 20 µg

Lanes 4, 8 and 12:

Jurkat (human T cell leukemia cell line from peripheral blood) 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 1/20000 dilution

Predicted band size: 98 kDa

Observed band size: 80 kDa,140 kDa

false

Exposure time: 180s