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Synthetic peptide conjugated to KLH derived from within residues 150 - 250 of Human macroH2A.1.
(Peptide available as ab37263.)
Our Abpromise guarantee covers the use of ab37264 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ChIP||Use at an assay dependent concentration. PubMed: 19380460|
|Flow Cyt||Use at an assay dependent concentration. ab171870-Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.|
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 40 kDa (predicted molecular weight: 40 kDa).Can be blocked with Rat macroH2A.1 peptide (ab37263).|
|IHC-P||Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
|ICC/IF||Use a concentration of 1 µg/ml.|
Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: MacroH2A.1 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: Hepg2 whole cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab37264 observed at 40 kDa. Red - loading control, ab18058, observed at 130 kDa.
ab37264 was shown to recognize macroH2A.1 in wild type cells as signal was lost at the expected MW in macroH2A.1 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and macroH2A.1 knockout samples were subjected to SDS-PAGE. Ab37264 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1 ug/ml and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.
ICC/IF image of ab37264 stained human MCF7 cells. The cells were methanol fixed (5 min), permabilised in 0.1% PBS-Tween (20 min) and incubated with the antibody (ab37264, 5µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue). This antibody also gave a positive IF result in HeLa and HEK 293 cells.
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"