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Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Flotillin 1.
(Peptide available as ab41926.)
Our Abpromise guarantee covers the use of ab41927 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|IHC-P||1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
|ICC/IF||1/200. Fix cells with methanol|
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 47 kDa (predicted molecular weight: 47 kDa).Can be blocked with Human Flotillin 1 peptide (ab41926).|
|IP||Use at an assay dependent concentration.|
Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: Flotillin 1 knockout HAP1 whole cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab41927 observed at 47 kDa. Red - loading control, ab9484, observed at 37 kDa.
ab41927 was shown to specifically react with Flotillin 1 in wild-type HAP1 cells as signal was lost in Flotillin 1 knockout cells. Wild-type and Flotillin 1 knockout samples were subjected to SDS-PAGE. ab41927 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1 µg/ml and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.
ICC/IF image of ab41927 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab41927 at 1µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
Immunohistochemical analysis of wholemount Xenopus laevis embryo, labelling Flotillin 1 with ab41927, diluted 1/50 and incubated in 10% Lamb Serum, 0.2% BSA, 0.1%TritonX/TBS for 14 hours at 4°C. A Goat Anti-Rabbit IgG (H+L) polyclonal secondary conjugated to Alexa Fluor® 555 was used, diluted 1/500. DAPI staining.
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