Anti-DNA2 antibody

• WB

Unknown

Western blot - Anti-DNA2 antibody (AB96488)

This antibody detects multiple bands in addition to the band of interest, reflecting low levels of expression of DNA2. siRNA knockdown experiments have demonstrated that this antibody detects a band at ~120 kDa corresponding to human DNA2 (please see image courtesy of Julien Duxin).

All lanes:

Western blot - Anti-DNA2 antibody (ab96488) at 1 µg/mL

Lane 1:

HEK293 (Human embryonic kidney cell line) Whole Cell Lysate at 10 µg

Lane 2:

HEK293 (Human embryonic kidney cell line) Nuclear Lysate at 10 µg

Lane 3:

U2OS (Human osteosarcoma cell line) Whole Cell Lysate at 10 µg

Secondary

All lanes:

Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Predicted band size: 120 kDa

Observed band size: 125 kDa,16 kDa,18 kDa,210 kDa,37 kDa,38 kDa,54 kDa,60 kDa,70 kDa,76 kDa

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Exposure time: 10min

• WB

Collaborator

Western blot - Anti-DNA2 antibody (AB96488)

Western blot showing detection of DNA2 by ab42439. A 120 kDa band corresponding to DNA2 was detected in lysates of U2OS cells treated with control shRNA (lane

1. but not in those from knockdown cells treated with two different shRNAs for DNA2 (lanes 2 and 3). Following pre-blocking for 30 mins in 5% milk, PBS-Tween, the membrane was incubated overnight at 4°C with ab42439 at a dilution of 1/1000 in 5% milk, PBS-Tween. The membrane was then incubated with the secondary antibody at 1/3000 dilution in 5% milk, PBS-Tween for 2 hours at RT, and washed 3 x 10 mins with PBS-Tween prior to ECL detection.

All lanes:

Western blot - Anti-DNA2 antibody (ab96488)

Lane 1:

U2OS cells plus control shRNA

Lane 2:

U2OS cells plus shDNA2-1 shRNA

Lane 3:

U2OS cells plus shDNA2-2 shRNA

Secondary

All lanes:

Anti-rabbit at 1/3000 dilution

Predicted band size: 120 kDa

Observed band size: 120 kDa

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Image courtesy of Julien Duxin and Sheila Stewart

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DNA2 antibody (AB96488)

IHC image of ab96488 staining in human liver carcinoma formalin fixed paraffin embedded tissue section, performed on a Leica Bond^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab96488, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

• WB

CiteAb

Western blot - Anti-DNA2 antibody (AB96488)

DNA2 western blot using anti-DNA2 antibody ab96488. Publication image and figure legend from Iannascoli, C., Palermo, V., et al., 2015, Nucleic Acids Res, PubMed 26275776.

ab96488 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab96488 please see the product overview.

Different exonucleases affect fork elongation in CPT, depending on the WRN status. (A) Western immunoblot showing depletion of DNA2 or EXO1 in cells expressing the wild-type form of WRN (WT) or its catalytically dead mutants (E84A and K577M). Whole cell extracts were prepared at 48 h after transfection or nucleofection with EXO1 siRNAs or DNA2 shRNA, respectively. The amount of EXO1 was evaluated after IP with the anti-EXO1 antibody. Lamin B1 was used as the loading control in WB with anti-DNA2 antibodies. l.e. = low exposure, h.e. = high exposure. Saturated signals are in red. (B) Analysis of IdU-labelled tract length of ongoing forks. The graphs show the mean value of IdU tract length (μm) from single DNA fibres in WS-derived cells stably expressing the WRN wild-type (WRN-WT) and the exonuclease-dead (WRN-E84A) or helicase-dead (WRN-K577M) mutants, in the presence (CPT) or absence (Untreated) of 50 nM CPT. Forty-eight hours before treatment, the cells were transfected with the indicated RNAi reagent. The length of the green tracks was measured in at least 100 well isolated DNA fibres from two independent experiments. Data are presented as mean ± SE (*p < 0.05; **p < 0.001 with respect to Ctrl RNAi; Kruskal–Wallis test). (C) Images of representative DNA fibres from the different cell lines.

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