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Synthetic peptide corresponding to Human Sortilin/NT3 aa 800 to the C-terminus (C terminal) conjugated to keyhole limpet haemocyanin.
(Peptide available as
Our Abpromise guarantee covers the use of ab16640 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 100 kDa (predicted molecular weight: 95 kDa).|
|ICC/IF||Use a concentration of 1 µg/ml.|
|Flow Cyt||Use at an assay dependent concentration. PubMed: 19761405
ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.
|IP||Use at an assay dependent concentration. PubMed: 21357693|
|IHC-FoFr||Use a concentration of 0.2 - 0.5 µg/ml.|
Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: Sortilin/NT3 knockout HAP1 cell lysate (20 µg)
Lane 3: NIH/3T3 cell lysate (20 µg)
Lane 4: 293T cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab16440 observed at 100 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab16640 was shown to specifically react with Sortilin/NT3 in wild-type HAP1 cells. No band was observed when Sortilin/NT3 knockout samples were examined. Wild-type and Sortilin/NT3 knockout samples were subjected to SDS-PAGE. ab16640 and ab8245 (loading control to GAPDH) were diluted 1 µg/mL and 1/10,000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.
Immunofluorescent staining for Sortilin/NT3 in rat cortical neurons using Rabbit polyclonal to Sortilin (ab16640). The staining is cytoplasmic and punctate of many cells such as cortical neurons. The image was taken with a X20 objective.
Protocol details: Rats were intracardially perfused with 4% paraformaldehyde. Whole brain tissue was post-fixed overnight in the same fixative, cryoprotected in 20% sucrose and frozen in OCT and then cut on cryostat (30µm coronal sections). IHC was perfored in free floating with fixed tissues (rat brain sections). Primary antibody was incubated overnight at 1/3000 at room temperature
ab16640 staining cultured mouse astrocytes by ICC/IF. The cultured cells were fixed with 4% paraformaldehyde for 5 minutes and blocked with 10% donkey serum in 0.1% PBS-0.3% TritonX for 30 minutes at 24°C. The cultured cells were then stained with ab16640 at 1/1000 in 0.3% TritonX with 0.1% PBS and 10% donkey serum for 4h at 24°C. An Alexa Fluro 488 donkey anti-rabbit polyclonal antibody at 1/1000 was used as the secondary antibody. Nuclei were stained with 1.43µM Hoechst and can be observed in blue. Sortilin expressed in the cytosol mainly in trans-golgi compartments.
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