Anti-APG5L/ATG5 antibody [EPR1755(2)] - BSA and Azide free

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-APG5L/ATG5 antibody [EPR1755(2)] - BSA and Azide free (AB221604)

Immunofluorescence staining of MCF7 cells with purified ab108327 at a working dilution of 1/150, counter-stained with DAPI. The secondary antibody was Alexa Fluor^® 488 goat anti-rabbit (ab150077), used at a dilution of 1/1000. ab7291, a mouse anti-tubulin antibody (1/1000), was used to stain tubulin along with ab150120 (Alexa Fluor^® 594 goat anti-mouse, 1/1000), shown in the top right hand panel. The cells were fixed in 100% methanol and permeabilized using 0.1% Triton X 100. The negative controls are shown in bottom middle and right hand panels - for negative control 1, purified ab108327 was used at a dilution of 1/500 followed by an Alexa Fluor^® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500. For negative control 2, ab7291 (mouse anti-tubulin) was used at a dilution of 1/500 followed by an Alexa Fluor^® 488 goat anti-rabbit antibody (ab150077) at a dilution of 1/400.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab108327).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] - BSA and Azide free (AB221604)

Unpurified ab108327, at 1/100 dilution, staining APG5L/ATG5 in paraffin-embedded Human hepatocellular carcinoma tissue by Immunohistochemistry.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab108327).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] - BSA and Azide free (AB221604)

Unpurified ab108327, at 1/100 dilution, staining APG5L/ATG5 in paraffin-embedded Human ovarian adenocarcinoma tissue by Immunohistochemistry.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab108327).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] - BSA and Azide free (AB221604)

Immunohistochemical staining of paraffin embedded human cervical carcinoma with purified ab180327 at a working dilution of 1/150. The secondary antibody used is HRP goat anti-rabbit IgG H&L (ab97051) at 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab108327).

• IP

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Immunoprecipitation - Anti-APG5L/ATG5 antibody [EPR1755(2)] - BSA and Azide free (AB221604)

ab180327 (purified) at 1/20 immunoprecipitating CRSP8 in 10 �g PC-12 whole cell lysate (Lanes 1 and 2, observed at 55 kDa). Lane 3 - PBS. For western blotting, VeriBlot for IP (HRP) (ab131366) was used as the secondary antibody (1/1500). Blocking buffer and concentration : 5% NFDM/TBST Dilution buffer and concentration : 5% NFDM/TBSTThis data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab108327).

All lanes:

Immunoprecipitation - Anti-APG5L/ATG5 antibody [EPR1755(2)] (<a href='/en-us/products/primary-antibodies/apg5l-atg5-antibody-epr17552-ab108327'>ab108327</a>)

Predicted band size: 32 kDa

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• WB

Lab

Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] - BSA and Azide free (AB221604)

False colour image of Western blot : Anti-APG5L/ATG5 antibody [EPR1755(2)] 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, ab108327 was shown to bind specifically to APG5L/ATG5. A band was observed at 50 kDa in wild-type THP-1 cell lysates with no signal observed at this size in ATG5 knockout cell line ab277835 (knockout cell lysate ab290722). To generate this image, wild-type and ATG5 knockout THP-1 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % 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-APG5L/ATG5 antibody [EPR1755(2)] (<a href='/en-us/products/primary-antibodies/apg5l-atg5-antibody-epr17552-ab108327'>ab108327</a>) at 1/1000 dilution

Lane 1:

Wild-type THP-1 cell lysate at 20 µg

Lane 2:

ATG5 knockout THP-1 cell lysate at 20 µg

Lane 3:

U-87 MG cell lysate at 20 µg

Lane 4:

HeLa cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Goat anti-Rabbit IgG H&L 800CW at 1/20000 dilution

Lanes 1 - 4:

Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Observed band size: 50 kDa

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