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Synthetic peptide derived from residues 200 to the C-terminus of Human HPRT.
Our Abpromise guarantee covers the use of ab10479 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/500 - 1/1000. Predicted molecular weight: 24 kDa.|
|ICC/IF||Use a concentration of 5 µg/ml.|
|IHC-P||1/200. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: HPRT1 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: A431 whole cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab10479 observed at 25 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab10479 was shown to specifically react with HPRT1 in wild-type HAP1 cells. No band was observed when HPRT1 knockout samples were examined. Wild-type and HPRT1 knockout samples were subjected to SDS-PAGE. Ab10479 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/500 dilution and 1/10,000 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 1/10,000 dilution for 1 hour at room temperature before imaging.
ICC/IF image of ab10479 stained human HeLa cells. The cells were methanol fixed ( 5 min) and incubated with the antibody (ab10479, 5µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 phalloidin was used to label F-actin (red).
Image courtesy of Human Protein Atlas. ab10479 staining HPRT in human small intestine. Paraffin embedded human small intestine tissue was incubated with ab10479 (1/200 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6.
ab10479 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines. Further results for this antibody can be found at www.proteinatlas.org