Huntington (phospho-S421) antibody

• Catalog Number: orb345421
• Category: Antibodies
• Description: Huntington (phospho-S421) antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: ELISA, IHC, WB
• Reactivity: Human
• Isotype: IgG
• Immunogen: Huntingtin pS421 Antibody was prepared from whole rabbit serum produced by repeated immunizations with a synthetic peptide corresponding to an internal region near aa 400-425 of Human Huntington Disease Protein.
• Concentration: 0.96 mg/mL
• Dilution range: ELISA: 1:10,000 - 1:40,000, IHC: 1:50 - 1:100, WB: 1:500 - 1:3,000
• Form/Appearance: Liquid (sterile filtered)
• Purity: Anti-Huntingtin pS421 is an affinity purified antibody produced by immunoaffinity chromatography using phospho peptide coupled to agarose beads followed by solid phase adsorption(s) against non-phospho peptide and non-specific peptide to remove any unwanted reactivities. This antibody is specific for phosphorylated human Huntington protein at the pS421 residue. BLAST analysis indicates 100 % homology of the immunizing sequence with Huntington homologues from chimpanzee, pig and chicken. Cross reactivity with Huntington protein homologues from mouse and rat may also occur as sequence homology varies by one amino acid residue in this sequence. Reactivity with Huntington protein from other sources is not known. Minimal reactivity is expected with the non-phosphorylated form of the protein.
• Conjugation: Unconjugated
• UniProt ID: P42858
• NCBI: NP_002102.4
• Storage: Store vial at -20° C prior to opening. Aliquot contents and freeze at -20° C or below for extended storage. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4° C as an undiluted liquid. Dilute only prior to immediate use.
• Buffer/Preservatives: 0.01% (w/v) Sodium Azide
• Alternative names: rabbit anti-Huntington pS421 antibody, rabbit anti
• Note: For research use only
• Application notes: Anti-Huntingtin pS421 antibody has been tested for use in ELISA, immunohistochemistry, and by western blot. Specific conditions for reactivity should be optimized by the end user. Expect bands at approximately 350 kDa and 200 kDa in size corresponding to full-length Huntingtin protein and truncated (hypothetical) Huntingtin protein, respectively, by western blotting in the appropriate cell lysate or extract. This antibody is specific for the phosphorylated form of Huntingtin protein at the pS421 residue. The identity of lower molecular bands ~130 kDa is not known.
• Expiration Date: 12 months from date of receipt.

Biorbyt's Affinity Purified anti-Huntingtin pS421 antibody was used at a 1:100 dilution to detect phosphorylated Huntingtin by immunohistochemistry in human brain cerebellum. Positive cytoplasmic staining is observed in neurons. Tissue was formalin-fixed and paraffin embedded.

Western blot analysis after AKT and phosphatase treatment is shown using Biorbyt's Affinity Purified anti-Huntingtin pS421 antibody. In A) untreated lysates from N2A and 293A cells were stained directly using anti-Huntingtin pS421 antibody. Full length staining of Huntingtin is noted, albeit at low levels of expression, as well as a strongly staining band at 200 kDa that may represent staining of truncated protein. In B) staining is shown after immunoprecipitation using a monoclonal antibody followed by AKT treatment (to phosphorylate), untreated, and phosphatase (PP) treated (dephosphorylate) immunoprecipitated HTT. Full length phosphorylated huntingtin is clearly detected in these immunopurified samples (except dephosphorylated). In C) lysates are treated directly with AKT or PP to alter the phosphorylation status of HTT.

Western blot analysis is shown using Biorbyt's Affinity Purified anti-Huntingtin pS421 antibody to detect endogenous protein present in an unstimulated human PC-3 whole cell lysate (arrowhead). Comparison to a molecular weight marker indicates a band of ~190 kDa corresponding to truncated human Huntingtin protein. The blot was incubated with a 1:1000 dilution of the antibody at room temperature followed by detection using standard techniques.