Anti-ISG15 antibody [EPR3446] - BSA and Azide free

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-ISG15 antibody [EPR3446] - BSA and Azide free (AB248489)

This data was developed using ab133346, the same antibody clone in a different buffer formulation.Immunofluorescent analysis of ISG15 in HeLa cells stained with ab133346 at a 1/100 dilution.

• WB

Lab

Western blot - Anti-ISG15 antibody [EPR3446] - BSA and Azide free (AB248489)

This data was developed using ab133346, the same antibody clone in a different buffer formulation.

Blocking and dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-ISG15 antibody [EPR3446] (<a href='/ko/products/primary-antibodies/isg15-antibody-epr3446-ab133346'>ab133346</a>) at 1/2000 dilution

Lane 1:

Untreated HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HeLa (Human cervix adenocarcinoma epithelial cell) treated with 10 ng/ml IFN- a1 for 16 hours whole cell 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 1/20000 dilution

Predicted band size: 18 kDa

Observed band size: 17 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-ISG15 antibody [EPR3446] - BSA and Azide free (AB248489)

This data was developed using ab133346, the same antibody clone in a different buffer formulation.

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)

Lane 2 : ISG15 IFN treated wild type HAP1 whole cell lysate (20 μg)

Lane 3 : ISG15 (KO) whole cell lysate (20 μg)

Lane 4 : ISG15 (KO) IFN alpha treated whole cell lysate (20 μg)

Lane 5 : Daudi whole cell lysate (20 μg)

Lanes 1 - 5 : Merged signal (red and green). Green - ab133346 observed at 18 kDa. Red - loading control, ab18058, observed at 130 kDa.

ab133346 was shown to specifically react with ISG15 in IFN alpha1 induced wild-type HAP1cells. Loss of signal was observed when knockout cell line was used. Wild-type and HAP1 IFN treated knockout samples were subjected to SDS-PAGE. ab133346 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1000 dilution and 1/30000 dilution respectively. Blots were developed 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 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-ISG15 antibody [EPR3446] (<a href='/ko/products/primary-antibodies/isg15-antibody-epr3446-ab133346'>ab133346</a>)

Predicted band size: 18 kDa

Observed band size: 18 kDa

false