Anti-LSD2 / AOF1 antibody [EPR18508]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A431 (Human epidermoid carcinoma) cells labeling LSD2 / AOF1 with ab193080 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on A431 cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

The negative controls are as follows : -
-ve control 1 : ab193080 at 1/500 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labelling LSD2 / AOF1 antibody (red) with purified ab193080 at a dilution of 1/70. Goat anti rabbit IgG (Alexa Fluor^® 488) was used as the secondary antibody at 1/2000. Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Isotype control antibody was Rabbit monoclonal IgG (black). The blue line shows cells without incubation with primary antibody and secondary antibody.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling LSD2 / AOF1 with ab193080 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on HeLa cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

The negative controls are as follows :
-ve control 1 : ab193080 at 1/500 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

LSD2 / AOF1 was immunoprecipitated from 1mg of K562 (Human chronic myelogenous leukemia cells from bone marrow) whole cell lysate with ab193080 at 1/80 dilution.

Lane 1 : K562 whole cell lysate 10ug (Input).

Lane 2 : ab193080 IP in K562 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab193080 in K562 whole cell lysate.

Western blot was performed from the immunoprecipitate using ab193080 at 1/1000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500.

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

Exposure time : 8 seconds.

All lanes:

Immunoprecipitation - Anti-LSD2 / AOF1 antibody [EPR18508] (ab193080)

Predicted band size: 92 kDa

false

• WB

Lab

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Lane 1 : Wild-type HAP1 cell lysate (20 μg)
Lane 2 : LSD2 / AOF1 knockout HAP1 cell lysate (20 μg)
Lane 3 : PC12 cell lysate (20 μg)
Lane 4 : Raw264.7 cell lysate (20 μg)
Lanes 1 - 4 : Merged signal (red and green). Green - ab193080 observed at 95 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab193080 was shown to specifically react with LSD2 / AOF1 when LSD2 / AOF1 knockout samples were used. Wild-type and LSD2 / AOF1 knockout samples were subjected to SDS-PAGE. ab193080 and ab8245 (loading control to GAPDH) were diluted at 1/2000 and 1/10 000 respectively and incubated overnight at 4°C. 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 1/10 000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (ab193080)

Predicted band size: 92 kDa

false

• WB

Supplier Data

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (ab193080) at 1/2000 dilution

Lane 1:

Human fetal brain lysate at 10 µg

Lane 2:

Human fetal heart lysate at 10 µg

Secondary

All lanes:

Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 92 kDa

Observed band size: 92 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Lanes 1-4 : Merged signal (red and green). Green - ab193080 observed at 95 kDa. Red - loading control ab8245 observed at 36 kDa.

ab193080 Anti-LSD2 / AOF1 antibody [EPR18508] was shown to specifically react with LSD2 / AOF1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265969 (knockout cell lysate ab258016) was used. Wild-type and LSD2 / AOF1 knockout samples were subjected to SDS-PAGE. ab193080 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2. 5 hours 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.

All lanes:

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (ab193080) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human KDM1B (LSD2 / AOF1) knockout HeLa cell lysate (<a href='/ko/products/cell-lysates/human-kdm1b-lsd2-aof1-knockout-hela-cell-lysate-ab258016'>ab258016</a>) at 20 µg

Lane 3:

HAP1 cell lysate at 20 µg

Lane 4:

HCT116 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 92 kDa

Observed band size: 95 kDa

false

• WB

Supplier Data

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (ab193080) at 1/2000 dilution

Lane 1:

NIH/3T3 (Mouse embyro fibroblast cells) cell lysate at 20 µg

Lane 2:

Mouse thymus lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/50000 dilution

Predicted band size: 92 kDa

Observed band size: 92 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (AB193080)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-LSD2 / AOF1 antibody [EPR18508] (ab193080) at 1/2000 dilution

Lane 1:

C6 (Rat glial tumor cells) cell lysate at 10 µg

Lane 2:

RAW 264.7 (Mouse macrophage cells transformed with Abelson murine leukemia virus) cell lysate at 10 µg

Lane 3:

PC-12 (Rat adrenal gland pheochromocytoma) cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/1000 dilution

Predicted band size: 92 kDa

Observed band size: 92 kDa

false

Exposure time: 3min