CDC27 (phospho-T244) antibody

• Catalog Number: orb345515
• Category: Antibodies
• Description: CDC27 (phospho-T244) antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: DOT, ELISA, IP, WB
• Reactivity: Human
• Isotype: IgG
• Immunogen: This affinity purified antibody was prepared from whole rabbit serum produced by repeated immunizations with a synthetic peptide corresponding to an internal portion surrounding T244 of Human CDC27.
• Concentration: 1.0 mg/mL
• Dilution range: ELISA: 1:8,000 - 1:30,000, IP: 1:100, WB: 1:300 - 1:2,000
• Form/Appearance: Liquid (sterile filtered)
• Purity: This affinity purified antibody is directed against the phosphorylated form of human CDC27 at the pT244 residue. The product was affinity purified from monospecific antiserum by immunoaffinity purification. Antiserum was first purified against the phosphorylated form of the immunizing peptide. The resultant affinity purified antibody was then cross adsorbed against the non-phosphorylated form of the immunizing peptide. Reactivity occurs against human CDC27 pT244 protein and the antibody is specific for the phosphorylated form of the protein. Reactivity with non-phosphorylated human CDC27 is minimal by ELISA. The antibody does not cross-react with CDC27 phosphorylated at other sites. A BLAST analysis was used to suggest reactivity with this protein from human, rat, dog, bovine and chimpanzee based on 100% homology for the immunogen sequence. Cross reactivity with CDC27 protein from mouse, rat and chicken may occur as sequence homology varies by one amino acid residue in this sequence (90% homology). Cross reactivity with CDC27 homologues from other sources has not been determined.
• Conjugation: Unconjugated
• UniProt ID: P30260
• NCBI: 167466175
• 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-CDC27 pT244 Antibody, CDC 27, CDC-27,
• Note: For research use only
• Application notes: This affinity purified antibody has been tested for use in ELISA and by western blot. Specific conditions for reactivity should be optimized by the end user. Expect a band approximately 92 kDa in size corresponding to CDC27 by western blotting in the appropriate cell lysate or extract. Less than 1% reactivity is observed against the non-phosphorylated form of the immunizing peptide. This antibody is phospho specific for pT244 of CDC27.
• Expiration Date: 12 months from date of receipt.

Dot Blot of Rabbit Anti-CDC27pT244 Antibody. Dilutions in Columns: (1) 100 ng, (2) 33.33 ng, (3) 11.11 ng, (4) 3.7 ng, (5) 1.23 ng. Tested BSA Peptide Reactivity in Rows: (A) AKT1-BSA, (B) AKT1 pT308-BSA, (C) AKT1 S473-BSA, (D) AKT1 pS473-BSA, (E) CDC27 T244-BSA, (F) CDC27 pT244-BSA, (G) BSA control. Primary Antibody: Anti-CDC27pT244 at 1 µg/mL overnight at 2-8°C. Secondary Antibody: Goat anti-Rabbit IgG HRP (p/n orb347654) at 1:70000 at RT for 30 mins. Block: BlockOut Buffer (p/n orb348644).

Western blot using Biorbyt's Affinity Purified anti-CDC27 pT244 antibody shows detection of a band ~92 kDa corresponding to phosphorylated human CDC27 (arrowhead). Lane 1 shows lysate from asynchronous cells. Lane 2 shows lysate from cells treated with nocodazole. Phosphorylated CDC27 is mostly present only in cell preparations arrested in mitosis. Each lane contains approximately 30 µg of HeLa whole cell lysates separated by 12.5% SDS-PAGE followed by transfer to nitrocellulose. After blocking with 5% non-fat dry milk in TTBS, the membrane was probed with the primary antibody diluted to 1:500 for 1 h at room temperature followed by washes and reaction with a 1:5000 dilution of HRP Gt-a-Rabbit IgG [H&L] MX (orb347654) for 45 min at room temperature. ECL reagent was used for detection. Other detection systems will yield similar results. Data contributed by Bing Li, UT Southwestern.