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Our Abpromise guarantee covers the use of ab214 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use 1-2µg for 106 cells.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|ICC/IF||1/200 - 1/500. Wash cells with PBS, then wash with PBS containing 0.5% Triton X-100 and fix for 5 min with PBS containing 3% paraformaldehyde. Block cells for 1 h in PBS containing 15% fetal bovine serum (see Robinson et al).|
|WB||Use a concentration of 1 - 5 µg/ml. Detects a band of approximately 79 kDa (predicted molecular weight: 79 kDa). (see Robinson et al).|
|IP||Use a concentration of 1 - 5 µg/ml. (for normal lymphoblastoid cell lines).|
|IHC-P||Use at an assay dependent concentration.|
Western blot of PC12 (brain)whole cell extracts at 30µg per lane, labeling Mre11 with ab214 at 1:500. A HRP-conjugated anti-mouse IgG antibody was used to detect the primary antibody.
ab214 staining Mre11 in human normal and gastric cancer tissue sections (20µm) by Immunohistochemistry (IHC-Fr - frozen sections). Tissue was fixed with paraformaldehyde for 3 hours, permeabilized with 0.1% Triton X-100 and blocked with 5% serum for 1 hour at 23°C. Samples were incubated with primary antibody (1/200 in 3% BSA + 0.1% Triton X-100 in TBS buffer) for 12 hours at 4°C. An undiluted HRP-conjugated goat anti-mouse IgG polyclonal was used as the secondary antibody.
Indirect immunofluorescence to detect localisation of Mre11 in normal lymphoblastoid cells.
(Panel D - negative control).
This picture was kindly supplied as part of the reviews for ab214 and for ab89 submitted by Anya Polischouk.
The bands represent nuclear (N) and cytoplasmic (C) extract from human lung cancer cells (U1810).
Overlay histogram showing HeLa cells stained with ab214 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab214, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was a goat anti-mouse DyLight® 488 (IgG, H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
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