Human LAMP2 knockout HeLa cell lysate

• WB

Unknown

Western blot - Human LAMP2 knockout HeLa cell lysate (AB263861)

Lane 1 : Wild-type HEK-293 cell lysate (20 μg)

Lane 2 : LAMP2 knockout HEK-293 cell lysate (20 μg)

Lane 3 : Wild-type HeLa cell lysate (20 μg)

Lane 4 : LAMP2 knockout HeLa cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab199946 observed at 110 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab199946 was shown to react with LAMP2 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab255402 (knockout cell lysate ab263861) was used. Wild-type and LAMP2 knockout samples were subjected to SDS-PAGE. ab199946 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 μg/ml and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-LAMP2 antibody [EPR19512] - Lysosome Marker (<a href='/en-us/products/primary-antibodies/lamp2-antibody-epr19512-lysosome-marker-ab199946'>ab199946</a>) at 1 µg/mL

Lane 1:

Wild-type HEK-293 cell lysate at 20 µg

Lane 2:

LAMP2 knockout HEK-293 cell lysate at 20 µg

Lane 3:

Wild-type HeLa cell lysate at 20 µg

Lane 4:

LAMP2 knockout HeLa cell lysate at 20 µg

Predicted band size: 45 kDa

Observed band size: 110 kDa,37 kDa

false

• WB

Lab

Western blot - Human LAMP2 knockout HeLa cell lysate (AB263861)

False colour image of Western blot : Anti-LAMP2A antibody [EPR4207(2)] - Lysosome Marker staining at 1/2000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab125068 was shown to bind specifically to LAMP2A. A band was observed at 100 kDa in wild-type HeLa cell lysates with no signal observed at this size in LAMP2 CRISPR-Cas9 edited cell line ab255402 (CRISPR-Cas9 edited cell lysate ab263861). The band observed in the CRISPR-Cas9 edited lysate lane below 100 kDa is likely to represent a truncated form of LAMP2A. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and LAMP2 CRISPR-Cas9 edited HeLa 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-LAMP2A antibody [EPR4207(2)] - Lysosome Marker (<a href='/en-us/products/primary-antibodies/lamp2a-antibody-epr42072-lysosome-marker-ab125068'>ab125068</a>) at 1/2000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

LAMP2 CRISPR-Cas9 edited HeLa 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: 100 kDa

false

• WB

Lab

Western blot - Human LAMP2 knockout HeLa cell lysate (AB263861)

False colour image of Western blot : Anti-LAMP2 antibody [EPR19531] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab199947 was shown to bind specifically to LAMP2. A band was observed at 100 kDa in wild-type HeLa cell lysates with no signal observed at this size in LAMP2 CRISPR-Cas9 edited cell line ab255402 (CRISPR-Cas9 edited cell lysate ab263861). The band observed in the CRISPR-Cas9 edited lysate lane below 100 kDa is likely to represent a truncated form of LAMP2. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and LAMP2 CRISPR-Cas9 edited HeLa 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-LAMP2 antibody [EPR19531] - Lysosome Marker (<a href='/en-us/products/primary-antibodies/lamp2-antibody-epr19531-lysosome-marker-ab199947'>ab199947</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

LAMP2 CRISPR-Cas9 edited HeLa 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: 100 kDa

false

• WB

Supplier Data

Western blot - Human LAMP2 knockout HeLa cell lysate (AB263861)

Lane 1 : Wild-type HEK-293 cell lysate (20 μg)

Lane 2 : LAMP2 knockout HEK-293 cell lysate (20 μg)

Lane 3 : Wild-type HeLa cell lysate (20 μg)

Lane 4 : LAMP2 knockout HeLa cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab199947 observed at 110 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab199947 was shown to react with LAMP2 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab255402 (knockout cell lysate ab263861) was used. Wild-type and LAMP2 knockout samples were subjected to SDS-PAGE. ab199947 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

false

• Sanger seq

Unknown

Sanger Sequencing - Human LAMP2 knockout HeLa cell lysate (AB263861)

Homozygous : Insertion of the selection cassette in exon 1