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Protein phosphatase associates with several regulatory proteins to produce holoenzymes required for removing phosphate groups from different targets. Protein phosphatase 1 (PP1) plays a large role in glycogen metabolism, protein synthesis and cell division. The Phospho-PP1alpha (T320) polyclonal antibody detects endogenous levels of PP1alpha protein only when phosphorylated at T320.
1. HOW DID OUR ANTIBODY PERFORM? ANTIBODY CUSTOMER REVIEW: Phospho-PP1alpha (T320) Polyclonal Antibody (STJ91134)
2. What was used? Primary antibody: STJ91134 Phospho-PP1alpha (T320) Provider: St John’s Laboratory Dilution ratio: 1:500 Application: Western Blot Materials for validation: HEK cell line Click for product data sheet PDF
3. What Was The Protocol? Treatment of Materials Cells were cultured to full confluence plating density. Lysis occurred with protease inhibitors (Roche), 1% triton, 50 mM Tris-HCL, 150 mM NaCl and a phosphatase inhibitors cocktail (Sigma). 5X of sample buffer was then added. Gel Electrophoresis 15% polyacrylamide gel, constant voltage 160 V, for 60 minutes. Transfer 0.45 um PVDF membrane. 1 piece of gel (semi-dry transfer), constant current 100V, for 60 minutes. Blocking 1X TBST with 5% BSA for 1 hour. Primary Antibody Probing Primary antibody was added to the solution of 1X TBST with 5% BSA, 4?C and left overnight. Membrane Wash Membrane was washed with 1X TBST 3 times. Secondary Antibody Probing Secondary antibody was added to the solution of 1X TBST with 5% BSA with a 1:2000 dilution ratio, for 1.5 hours. Membrane Wash Membrane was washed with 1X TBST 3 times. Visualization Visualization: ECL visualization for 3 min (the visualization time was determined by actual result. It was ensured that the visualization time was identical for the same antigen).
4. Gel electrophoresis information: 15% polyacrylamide gel, constant voltage 160 V, 60 min. Transfer information: 0.45 nm PVDF membrane, 1 piece of gel (wet transfer), constant voltage 100 V 60 min. No. Antigen Loading amount Primary antibody Primary antibody dilution ratio Secondary antibody dilution ratio Target band KD Visualization time 1 Hek lysate 40ug STJ91134 phospho PP1 1:500 1:2000 ~37KD 60S
5. What were the results?
6. What did the customer think? Antibody Specificity: Antibody Rating: Testing results were provided independently by S. Hilfiker at Instituto de Parasitolog¨ªa y Biomedicina “L¨®pez – Neyra”.
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