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Synthetic peptide derived from within residues 200 - 300 of Human Cathepsin B.
Synthetic peptide of Human Cathepsin B.
Our Abpromise guarantee covers the use of ab92955 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use a concentration of 1 µg/ml.|
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 30, 25 kDa (predicted molecular weight: 38 kDa).|
Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: Cathepsin B knockout HAP1 whole cell lysate (20 µg)
Lane 3: A549 whole cell lysate (20 µg)
Lane 4: HL60 whole cell lysate (Negative Control) (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab92955 observed at 24-27-44 kDa. Red - loading control, ab18058, observed at 130 kDa.
ab92955 was shown to recognize Cathepsin B in wild-type HAP1 cells as signal was lost at the expected MW in Cathepsin B knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and Cathepsin B knockout samples were subjected to SDS-PAGE. ab92955 and ab18058 (Mouse anti-Vinculin 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.
Pro-cathepsin B runs at approximately 44 kDa on SDS-PAGE. When it is proteolytically processed and glycosylated it forms a mature two-chain protein containing a heavy chain (running at 27 and 24 kDa) and a light chain (5 kDa)
ICC/IF image of ab92955 stained HepG2 cells. The cells were 100% methanol fixed (5 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 ab92955 at 1µg/ml overnight at +4°C. 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) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
ab92955 has not yet been referenced specifically in any publications.
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