Recombinant Anti-ATM antibody [Y170] (ab32420)

False colour image of Western blot: Anti-ATM antibody [Y170] 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, ab32420 was shown to bind specifically to ATM. A band was observed at 350 kDa in wild-type A549 cell lysates with no signal observed at this size in ATM knockout cell line ab276095 (knockout cell lysate ab283834). To generate this image, wild-type and ATM 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 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 (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

Immunocytochemistry/ Immunofluorescence analysis of HeLa cells labeling ATM with ab32420 at 1/500. Goat anti rabbit IgG(Alexa Fluor^® 488), ab150077 at 1/1000 was used as the secondary antibody. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% tritonX-100. Nulei were counterstained blue with DAPI.

Immunocytochemistry/ Immunofluorescence analysis of HepG2 cells labeling ATM with ab32420 at 1/500. Goat anti rabbit IgG(Alexa Fluor^® 488), ab150077 at 1/1000 was used as the secondary antibody. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% tritonX-100. Nulei were counterstained blue with DAPI.

DNA repair proteins accumulate at MVM APAR bodies

Repair proteins accumulate at APAR bodies. NB324K cells were infected with MVMp (MOI of 10) for 16 hr before being fixed and processed for immunofluorescence. Cells were stained with the indicated antibodies to mark DDR repair proteins. APAR bodies were detected with antibodies to NS1. Nuclei were stained with DAPI. All images were captured using an objective of 63×. 

Cells were fixed with 4% paraformaldehyde for 15 minutes and permeabilized with 0.5% Triton X-100 in PBS for 15 minutes.

(Image shows the right-hand panel of Figure 2A)

Formaldehyde-fixed human colon tissue stained for ATM using ab32420 at 1/100 dilution in immunohistochemical analysis.

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Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labelling ATM with purified ab32420 at 1/20 (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. An Alexa Fluorr^®488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies. 

Immunohistochemical analysis of paraffin-embedded Human breast cancer tissue labeling ATM with ab32420 at 1:100 dilution. Tissue underwent antigen retrieval using Tris/EDTA Buffer (pH9.0). The section was counterstained with haematoxylin.

Formaldehyde-fixed human serous ovarian tumor tissue stained for ATM using ab32420 at 1/50 dilution in immunohistochemical analysis.

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Immunohistochemical analysis of paraffin-embedded Human normal breast tissue labeling ATM with ab32420 at 1:100 dilution. Tissue underwent antigen retrieval using Tris/EDTA Buffer (pH9.0). The section was counterstained with haematoxylin.