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Recombinant full length protein (Human).
Please note that this antibody is not suitable for WB.
Despite positive publications and Abreviews we have mixed feedback on this antibody in WB and we do not guarantee ab16780 for WB.
Our Abpromise guarantee covers the use of ab16780 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 1µg for 106 cells.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|ICC/IF||Use at an assay dependent concentration.|
|IHC-Fr||Use at an assay dependent concentration.|
|IP||Use at an assay dependent concentration.|
|IHC-P||Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.|
ICC/IF image of ab16780 stained MCF7cells. The cells were 4% PFA fixed (10 minutes) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1 hour to permeabilize the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab16780, 5 µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-mouse IgG - H&L, pre-adsorbed (ab96879) used at a 1/250 dilution for 1 hour. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1 hour. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43 µM.
Overlay histogram showing MCF7 cells stained with ab16780 (red line). The cells were fixed with 80% methanol (5 minutes) and then permeabilized with 0.1% PBS-Tween for 20 minutes. 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 (ab16780, 1 µg/1x106 cells) for 30 minutes at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 minutes at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2 µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in MCF7 cells fixed with 4% paraformaldehyde (10 minutes)/permeabilized with 0.1% PBS-Tween for 20 minutes used under the same conditions.
Immunoprecipitation step: Protein G
Sample: HeLa whole cell lysate
Total protein input: 2000 µg
Concentration: 1 µg/mg lysate
Incubation time: 2 hours at 4°C
WB antibody: Rabbit polyclonal, HRP conjugate at 1/2500 dilution
ab16780 staining BRAC1 in human ovarian tumor cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with Acetone:Methanol and blocked with a protein block, serum-free for 1 hour at 18°C. Samples were incubated with primary antibody (1/100) for 14 hours at 4°C. An Alexa Fluor® 488-conjugated Rabbit anti-mouse IgG (H+L) polyclonal (1/400) was used as the secondary antibody.
ab16780 staining BRCA1 in human skin tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded tissue sections). The sections were fixed in formaldehyde and subjected to heat-mediated antigen retrieval in citrate buffer (pH 6.0) prior to blocking with 2% BSA for 1 hour at 22°C. The primary antibody was diluted 1/50 and incubated with the sample for 20 hours at 4°C. A biotin-conjugated goat anti-mouse polyclonal was used as the secondary antibody, diluted 1/800. Antibody was detected by DAB staining.
ab16780 staining BRAC1 in human colon cancer cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.3% Triton X-100 and blocked with 3% BSA for 30 minutes at 4°C. Samples were incubated with primary antibody (1/200) for 12 hours at 4°C. An Alexa Fluor® 488-conjugated Goat anti-mouse IgG (H+L) polyclonal (1/1000) was used as the secondary antibody.
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"