Pituitary Tumor-Transforming 1 (PTTG1) antibody

Antigen: Pituitary Tumor-Transforming 1 (PTTG1)

Synonyms: EAP1, PTTG, HPTTG, TUTR1, MGC126883, MGC138276, PTTG1, MGC127932, MGC134134

Clonality: Monoclonal (DCS-280)

Host: Mouse

Reactivity: Human

Application: Immunofluorescence (IF), Immunoprecipitation (IP), Immunohistochemistry (Paraffin-embedded Sections) (IHC (p)), Western Blotting (WB)

Catalog no.: ABIN487310

Additional Information

Characteristics: Synonyms: PTTG, EAP1, TUTR1, Pituitary tumor-transforming gene 1 protein, Esp1-associated protein,Tumor-transforming protein 1

Alternative name: Securin / PTTG1

Gene ID: 9232

Swiss-Prot: O95997

Immunogen: Recombinant full-length Human Securin. Remarks: Hybridoma was established by fusion of Mouse myeloma cell NS-2 with Balb/cmouse splenocyte

Cross-Reactivity: Human, Mouse (Murine), Rat (Rattus)

Isotype: IgG2a  (Matching secondary antibodies)

Clone: DCS-280

Description: The anaphase inhibitor PDS1, also known as securin or pituitary tumor-transforming gene-1(PTTG1) protein, is required for chromosomal stability and for the maintenance of euploidyin human cells. Human cells without a PDS1 gene lose chromosomes at a high rate. Theselosses have been linked to abnormal anaphases during which cells undergo repeatedunsuccessful attempts to segregate their chromosomes. PDS1 binds to Ku, the regulatorysubunit of the DNA-dependent protein kinase, both in vitro and in vivo. DNA double-strandbreaks prevent Ku/PDS1 association, suggesting that genome damaging events can inducepathways that activate DNA repair mechanisms and halt cell cycle progression.

Specificity: This antibody reacts with Securin/Pds-1 (28 kDa) on Western blotting. Species Reactivity: Tested: Human, Mouse and Rat. Add. Information: This product was originally produced by MBL International.

Application Details

Protocol: SDS-PAGE & Western Blotting1) Wash the cells 3 times with PBS and suspend with 10 volume of cold Lysis buffer (50 mMTris-HCl, pH 7. 2, 250 mM NaCl, 0. 1% NP-40, 2 mM EDTA, 10% glycerol) containingappropriate protease inhibitors. Incubate it at 4°C with rotating for 30 minutes, thensonicate briefly (up to 10 seconds). 2) Centrifuge the tube at 12,000 x g for 10 minutes at 4°C and transfer the supernatant toanother tube. Measure the protein concentration of the supernatant and add the Lysisbuffer to make 8 mg/mL solution. 3) Mix the sample with equal volume of Laemmli’s sample buffer. 4) Boil the samples for 2 minutes and centrifuge. Load 10 μL of the sample per lane in a 1mm thick SDS-polyacrylamide gel for electrophoresis. 5) Blot the protein to a polyvinylidene difluoride (PVDF) membrane at 1 mA/cm2 for 1 hourin a semi-dry transfer system. (Transfer Buffer: 25 mM Tris, 190 mM glycine, 20% MeOH). See the manufacture's manual for the transfer procedure. 6) To reduce nonspecific binding, soak the membrane in 10% skimmed milk (in PBS, pH7. 2) for 1 hour at room temperature, or overnight at 4°C. 7) Incubate the membrane with primary antibody diluted with PBS, pH 7. 2 containing 1%skimmed milk as suggest in the APPLICATIONS for 1 hour at room temperature. (Theconcentration of antibody will depend on condition. )8) Wash the membrane with PBS (5 minutes x 6 times). 9) Incubate the membrane with the 1: 10000 POD-conjugated anti-mouse IgG diluted with1% skimmed milk (in PBS, pH 7. 2) for 1 hour at room temperature. 10) Wash the membrane with PBS (5 minutes x 6 times). 11) Wipe excess buffer from the membrane, then incubate it with appropriatechemiluminescence reagents for 1 minute. Remove extra reagent from the membrane bydabbing with a paper towel, and seal it in plastic wrap. 12) Expose to an X-ray film in a dark room for 5 minutes. Develop the film as usual. Theconditions for exposure and development may vary. Positive Controls for Western blotting: Raji, Jurkat, U251, A431, WR19L, C2C12, Rat-1. Immunoprecipitation1) Wash the cells 3 times with PBS and suspend with 10 volume of cold Lysis buffer (50 mMTris-HCl, pH 7. 2, 250 mM NaCl, 0. 1% NP-40, 2 mM EDTA, 10% glycerol) containingappropriate protease inhibitors. Incubate it at 4°C with rotating for 30 minutes, then

Application Notes: Western Blot: 0.2-1.0 µg/ml for chemiluminescence detection systemPositive Controls: Human: Raji, Jurkat, U251, A431. Mouse: WR19L, C2C12. Rat: Rat-1Immunoprecipitation: 2 µg/200 µL of cell extract from 5x10e6 cellsPositive Control: Raji. Immunohistochemistry: 1-5 μg/mLHeat treatment is necessary for Paraffin Embedded Sections. Microwave oven: 2 times for 10 minutes each in citrate buffer (pH 6.5). Positive Control: Tonsil Tissue.

Concentration: 1.0 mg/ml

Purification: Azide Free

Buffer: PBS, pH 7.2 containing 50% Glycerol without preservatives.

Storage: Store the antibody (in aliquots) at -20°C. Avoid repeated freezing and thawing. Shelf life: one year from despatch.

Restrictions: For Research Use only