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569388 Anti-STAT3 Rabbit pAb

Anti-STAT3 Rabbit pAb: SDB (Sicherheitsdatenblätter), Analysenzertifikate und Qualitätszertifikate, Dossiers, Broschüren und andere verfügbare Dokumente.

Synonyme: Anti-Signal Transducer and Activator of Transcription 3

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      Übersicht

      Replacement Information

      Key Spec Table

      Host
      Rb
      Description
      Overview

      This product has been discontinued.



      We are offering Anti-Stat3 Antibody (Cat. No. 06-596) as a possible alternative. Please read the alternative product documentation carefully and contact technical service if you need additional information.






      Recognizes native and denatured STAT3.
      Catalogue Number569388
      Brand Family Calbiochem®
      SynonymsAnti-Signal Transducer and Activator of Transcription 3
      Application Data
      Detection of human STAT3 by immunoblotting. Samples: Whole cell lysate from HeLa cell treated with IFN-α (lane 1) or left untreated (lane 2). Primary antibody: Anti-STAT3 Rabbit pAb (Cat. No. 569388) (1:1000). Detection: chemiluminescence.
      References
      ReferencesDavid, M., et al. 1995. Science 269, 1721.
      Ihle, J.N. 1995. Nature 377, 591.
      Wen, Z., et al. 1995. Cell 82, 241.
      Darnell, J.E., Jr., et al. 1994. Science 264, 1415.
      Ihle, J.N., et al. 1994. Trends Biochem. Sci. 19, 222.
      Product Information
      FormLiquid
      FormulationIn 150 mM NaCl, 10 mM HEPES, 50% glycerol, 0.01% BSA, pH 7.5.
      PreservativeNone
      Quality LevelMQ100
      Applications
      Key Applications Immunoblotting (Western Blotting)
      Immunoprecipitation
      Paraffin Sections
      Application NotesImmunoblotting (1:1000)
      Paraffin Sections (1:100, heat pre-treatment required)
      Immunoprecipitation (1:100)
      Application CommentsFor paraffin sections, pre-treatment in citrate buffer, pH 6.0 is required. Detects total STAT3 (phosphorylation-state independent) levels. Recognizes both native and denatured STAT3. Variables associated with assay conditions will dictate the proper working dilution.

      Recommended Protocol for Immunoblotting

      Solutions and Reagents
      • Transfer Buffer: 25 mM Tris base, 0.2 M glycine, 20% methanol, pH 8.5.
      • SDS Sample Buffer: 62.5 mM Tris-HCl, pH 6.8, 2% SDS, 10% glycerol, 50 mM DTT, 0.1% bromphenol blue.
      • 10X TBS (Tris-buffered saline): To prepare 1 liter, 24.2 g Tris base, 80 g NaCl, adjust pH to 7.6 with HCl. Dilute 1:10 for use.
      • Blocking Buffer: 1X TBS, 0.1% Tween®-20 detergent with 5% non-fat dry milk.
      • Primary Antibody Dilution Buffer: 1X TBS, 0.1% Tween®-20 detergent with 5% BSA
      • Wash Buffer (TBST): 1X TBS, 0.1% Tween®-20 detergent

      Blotting Membrane
      Nitrocellulose or PVDF membranes may be used.

      Protein Blotting
      A general protocol for sample preparation using 2x106 SK-N-MC cells per well in a 6-well plate is as follows:

      1. Culture cells in medium containing 0.5% FBS for 2 days. We recommend plating cells directly in 0.5% FBS media to reduce basal levels of STAT3 phosphorylation.
      2. Aspirate media. Add fresh media without FBS. Culture for 2 h. Note: If cells are grown at high density, changing media before treating cells with regulator reduces basal STAT3 phosphorylation due to factors secreted by cells.
      3. Aspirate media. Treat cells by adding fresh media containing regulator for desired time.
      4. Aspirate media from cultures; wash cells with PBS; aspirate.
      5. Lyse cells by adding 100 µl SDS Sample Buffer and immediately scrape the cells off the plate and transfer the extract to a microfuge tube. Keep on ice.
      6. Sonicate for 2 s to shear DNA and reduce sample viscosity.
      7. Heat sample to 95-100°C for 5 min. Cool on ice.
      8. Microcentrifuge for 5 min.
      9. Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
      10. Electrotransfer to nitrocellulose membrane.

      As controls, we recommend using 10 µl of SK-N-MC cell extracts.

      Membrane Blocking, Gel and Antibody Incubations
      1. After transfer, wash membrane with 25 ml TBS for 5 min at room temperature.
      2. Incubate membrane in 25 ml Blocking Buffer for 1-3 h at room temperature or overnight at 4°C.
      3. Wash 3 times for 5 min each with 15 ml TBST.
      4. Incubate membrane and primary antibody (at the appropriate dilution) in 10 ml Primary Antibody Dilution Buffer with gentle agitation overnight at 4°C.
      5. Wash 3 times for 5 min each with 15 ml TBST.
      6. Incubate membrane with conjugated secondary antibody at the appropriate dilution in 10 ml Blocking Buffer with gentle agitation for 1 h at room temperature.
      7. Wash membrane as in step 5.

      Detection of Proteins
      Chemiluminescence.
      Biological Information
      Immunogena synthetic peptide corresponding to amino acids surrounding Tyr⁷⁰⁵ of mouse STAT3
      ImmunogenMouse
      HostRabbit
      IsotypeIgG
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Storage and Shipping Information
      Ship Code Blue Ice Only
      Toxicity Standard Handling
      Storage -20°C
      Avoid freeze/thaw Avoid freeze/thaw
      Do not freeze Ok to freeze
      Special InstructionsFollowing initial thaw, aliquot and freeze (-20°C).
      Packaging Information
      Transport Information
      Supplemental Information
      Specifications
      Global Trade Item Number
      Bestellnummer GTIN
      569388 0

      Documentation

      Anti-STAT3 Rabbit pAb Analysenzertifikate

      TitelChargennummer
      569388

      Literatur

      Übersicht
      David, M., et al. 1995. Science 269, 1721.
      Ihle, J.N. 1995. Nature 377, 591.
      Wen, Z., et al. 1995. Cell 82, 241.
      Darnell, J.E., Jr., et al. 1994. Science 264, 1415.
      Ihle, J.N., et al. 1994. Trends Biochem. Sci. 19, 222.
      Datenblatt

      Note that this data sheet is not lot-specific and is representative of the current specifications for this product. Please consult the vial label and the certificate of analysis for information on specific lots. Also note that shipping conditions may differ from storage conditions.

      Revision06-July-2010 JSW
      SynonymsAnti-Signal Transducer and Activator of Transcription 3
      ApplicationImmunoblotting (1:1000)
      Paraffin Sections (1:100, heat pre-treatment required)
      Immunoprecipitation (1:100)
      Application Data
      Detection of human STAT3 by immunoblotting. Samples: Whole cell lysate from HeLa cell treated with IFN-α (lane 1) or left untreated (lane 2). Primary antibody: Anti-STAT3 Rabbit pAb (Cat. No. 569388) (1:1000). Detection: chemiluminescence.
      DescriptionProtein A and immunoaffinity purified rabbit polyclonal antibody. Recognizes the ~90 kDa STAT3 protein.
      BackgroundSTAT3 is activated by a variety of cytokines including interleukins, CNTF and EPO. Activation of STAT3 is accompanied by tyrosine phosphorylation at Tyr705 which induces dimerization, nuclear translocation and DNA binding. Transcriptional activation also appears to be regulated by serine phosphorylation (Ser727) probably via MAP-like kinases. As phosphorylation of STAT3 at Tyr705 is essential for dimerization and DNA binding, phosphorylation at this site is an excellent marker of STAT3 activity.
      HostRabbit
      Immunogen speciesMouse
      Immunogena synthetic peptide corresponding to amino acids surrounding Tyr⁷⁰⁵ of mouse STAT3
      IsotypeIgG
      Specieshuman, mouse, rat
      FormLiquid
      FormulationIn 150 mM NaCl, 10 mM HEPES, 50% glycerol, 0.01% BSA, pH 7.5.
      PreservativeNone
      CommentsFor paraffin sections, pre-treatment in citrate buffer, pH 6.0 is required. Detects total STAT3 (phosphorylation-state independent) levels. Recognizes both native and denatured STAT3. Variables associated with assay conditions will dictate the proper working dilution.

      Recommended Protocol for Immunoblotting

      Solutions and Reagents
      • Transfer Buffer: 25 mM Tris base, 0.2 M glycine, 20% methanol, pH 8.5.
      • SDS Sample Buffer: 62.5 mM Tris-HCl, pH 6.8, 2% SDS, 10% glycerol, 50 mM DTT, 0.1% bromphenol blue.
      • 10X TBS (Tris-buffered saline): To prepare 1 liter, 24.2 g Tris base, 80 g NaCl, adjust pH to 7.6 with HCl. Dilute 1:10 for use.
      • Blocking Buffer: 1X TBS, 0.1% Tween®-20 detergent with 5% non-fat dry milk.
      • Primary Antibody Dilution Buffer: 1X TBS, 0.1% Tween®-20 detergent with 5% BSA
      • Wash Buffer (TBST): 1X TBS, 0.1% Tween®-20 detergent

      Blotting Membrane
      Nitrocellulose or PVDF membranes may be used.

      Protein Blotting
      A general protocol for sample preparation using 2x106 SK-N-MC cells per well in a 6-well plate is as follows:

      1. Culture cells in medium containing 0.5% FBS for 2 days. We recommend plating cells directly in 0.5% FBS media to reduce basal levels of STAT3 phosphorylation.
      2. Aspirate media. Add fresh media without FBS. Culture for 2 h. Note: If cells are grown at high density, changing media before treating cells with regulator reduces basal STAT3 phosphorylation due to factors secreted by cells.
      3. Aspirate media. Treat cells by adding fresh media containing regulator for desired time.
      4. Aspirate media from cultures; wash cells with PBS; aspirate.
      5. Lyse cells by adding 100 µl SDS Sample Buffer and immediately scrape the cells off the plate and transfer the extract to a microfuge tube. Keep on ice.
      6. Sonicate for 2 s to shear DNA and reduce sample viscosity.
      7. Heat sample to 95-100°C for 5 min. Cool on ice.
      8. Microcentrifuge for 5 min.
      9. Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
      10. Electrotransfer to nitrocellulose membrane.

      As controls, we recommend using 10 µl of SK-N-MC cell extracts.

      Membrane Blocking, Gel and Antibody Incubations
      1. After transfer, wash membrane with 25 ml TBS for 5 min at room temperature.
      2. Incubate membrane in 25 ml Blocking Buffer for 1-3 h at room temperature or overnight at 4°C.
      3. Wash 3 times for 5 min each with 15 ml TBST.
      4. Incubate membrane and primary antibody (at the appropriate dilution) in 10 ml Primary Antibody Dilution Buffer with gentle agitation overnight at 4°C.
      5. Wash 3 times for 5 min each with 15 ml TBST.
      6. Incubate membrane with conjugated secondary antibody at the appropriate dilution in 10 ml Blocking Buffer with gentle agitation for 1 h at room temperature.
      7. Wash membrane as in step 5.

      Detection of Proteins
      Chemiluminescence.
      Storage Avoid freeze/thaw
      -20°C
      Do Not Freeze Ok to freeze
      Special InstructionsFollowing initial thaw, aliquot and freeze (-20°C).
      Toxicity Standard Handling
      ReferencesDavid, M., et al. 1995. Science 269, 1721.
      Ihle, J.N. 1995. Nature 377, 591.
      Wen, Z., et al. 1995. Cell 82, 241.
      Darnell, J.E., Jr., et al. 1994. Science 264, 1415.
      Ihle, J.N., et al. 1994. Trends Biochem. Sci. 19, 222.