重组Anti-RAP80抗体[EPR5315]

• WB

Unknown

Western blot - Anti-RAP80 antibody [EPR5315] (AB124763)

All lanes:

Western blot - Anti-RAP80 antibody [EPR5315] (ab124763) at 1/10000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

Human ovary cancer lysate at 10 µg

Lane 3:

293T cell lysate at 10 µg

Lane 4:

HT-1080 cell lysate at 10 µg

Secondary

All lanes:

Goat anti-Rabbit HRP at 1/2000 dilution

Predicted band size: 80 kDa

false

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RAP80 antibody [EPR5315] (AB124763)

ab124763 at 1/250 staining RAP80 in paraffin-embedded Human testis tissue by Immunohistochemistry.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

• WB

CiteAb

Western blot - Anti-RAP80 antibody [EPR5315] (AB124763)

RAP80 western blot using anti-RAP80 antibody [EPR5315] ab124763. Publication image and figure legend from Renaud, E., Barascu, A., et al., 2016, Nucleic Acids Res, PubMed 26446986.

ab124763 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 ab124763 please see the product overview.

53BP1 and RAP80 altered foci accumulation in FA pathway-deficient cells. (A) Representative images of RAP80 (red) and 53BP1 (green) foci in nuclei (DAPI stained, blue) of FANCC-mutated (PD331 FANCC-/-) and -corrected (PD331corr) cells 24 h after MMC exposure (1 μg/ml/1 h). White line : 2 μm. (B) Representative images of RIF1 (red) and 53BP1 (green) foci in nuclei (DAPI stained, blue) of FANCC-mutated (PD331 FANCC-/-) and -corrected (PD331corr) cells 24 hours after MMC exposure (1 μg/ml/1 h). White line : 2 μm. (C) 53BP1 and RAP80 foci quantificationin untreatyed cells (0 h) and at different time points (1, 6 and 24 h) following MMC exposure (1 μg/ml/1 h). Histograms represent the mean of three independent experiments. Error bars indicate S.D. The data were analyzed by a Student's t-test; ** indicates p < 0.01. (D) Size (arbitrary unit, a.u.) of 53BP1 foci in untreated conditions and 24 h after MMC exposure (1 μg/ml/1 h) in FANCC-mutated (PD331 FANCC-/-) and -corrected (PD331corr) cells. The cell size was determined using the ImageJ software. The values on the histogram represent the mean of three independent experiments; at least 50 cells were scored each time. Error bars indicate S.D. Data were analyzed by a Student's t-test; *** indicates p < 0.001. (E) Western blot showing the efficiency of the siRNA against 53BP1 and RAP80, as observed 72 h after transfection, in corrected (PD331 corr) and FA-C (PD331 FANCC-/-) cells. Vinculin was used as a loading control. siCtrl (control) indicates cells transfected with an untargeted siRNA. (F) Representative images of RAP80 (red) and 53BP1 (green) foci in nuclei (DAPI stained, blue) of FANCC-corrected cells (PD331 corr) after depletion of 53BP1 or RAP80 by siRNA transfection. Cells were treated with MMC (1 μg/ml/1 h) and analyzed 24 h later. White line : 2 μm. siCtrl (control) indicates cells transfected with an untargeted siRNA. (G) Quantification of RAP80 foci-positive cells after transfection with an untargeted siRNA (siCtrl, control), si53BP1 or siMDC1 in FANCC-mutated (PD331 FANCC-/-) and -corrected (PD331corr). Cells presenting more than 5 foci were considered positive. Data represent the mean of two independent experiments with similar results. (H) Left. Representative images of BRCA1/RAP80 interaction dots detected with the Proximity Ligation Assay in nuclei (DAPI stained, blue) in FANCC-mutated (PD331 FANCC-/-) and -corrected (PD331corr) cells under untreated conditions or 24 h following MMC exposure (1 μg/ml/1 h). White line : 8 μm. Right. BRCA1/RAP80 interaction dots quantification 24 h after MMC exposure (1 μg/ml/1 h). Histograms represent the pooled data from three independent experiments. Quantification was conducted using ImageJ software. (I) Left. Representative images of BRCA1/CtIP (left) interaction dots detected with the Proximity Ligation Assay in nuclei (DAPI stained, blue) in FANCC-mutated (PD331 FANCC-/-) and -corrected (PD331corr) cells under untreated conditions or 24 hours following MMC exposure (1 μg/ml/1 h). White line : 8 μm. Right. BRCA1/CtIP interaction dots quantification 24 h after MMC exposure (1 μg/ml/1 h). Histograms represent the pooled data from three independent experiments. Quantification was conducted using ImageJ software. (J) Representative images of FANCD2 foci (red), RAP80 (red) or 53BP1 (green) in PD20 corr (FANCD2-corrected cells), PD20 (FANCD2-/- cells) and PD20 FANCD2 K561R (PD20 cells expressing a K561R, non-ubiquitinable FANCD2) cells. White line : 2 μm. Histograms represent the number of RAP80 (red) and 53BP1 (green) foci in the three different cell lines in the left. Data represent the mean of two independent experiments with similar results.

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