Western Blotting Sample Preparation Products

western blot prep

Introduction

Western blotting among the most frequently employed techniques used to detect proteins based on antigen-antibody interactions. The first step of western blotting is sample preparation; your protein of interest must be extracted from the tissue or cell culture it’s expressed in, protected from degradation, and prepared to be run on a gel. Boster provides products for sample preparation, including specialized extraction kits, lysis buffers, and protease inhibitors.

Procedure

To extract a protein sample from a cell culture on solid medium, first rinse the culture with ice-cold PBS solution, then add ice-cold lysis buffer. Scrape adherent cells off the dish using a cold plastic scraper, or digest the cells with 0.05% trypsin. Once the cells are suspended, centrifuge the sample, draw off the supernatant fluid, and wash the pellet with ice cold PBS buffer. Re-suspend the cells in Boster mammalian cell protein extraction reagent and sonicate the solution for about 10-15 seconds, then lyse the cells with Boster RIPA lysis buffer and additional sonication. Centrifuge the sample to separate lipids and cellular debris from the soluble cell proteins, and aliquot the middle layer from the centrifuged sample into a fresh tube and store it at -20C.

The aliquots of cell lysis supernatant are ready for total protein quantification and gel electrophoresis.

For a detailed protocol of sample preparation starting with tissue samples, see our Western blot handbook.

Tips

  1. For best results, select the right type extraction kit for your sample
  2. If you observe weak signal, ghost bands, or inconsistent band sizes in your results, the source of error may be the protein extraction step. See our troubleshooting guides for more information


Displaying 7 Items

    Plus RIPA Lysis Buffer (AR0102-100)

    • Specific References (1)
    • Q&As (2)
    SKU AR0102-100
    Product name Plus RIPA Lysis Buffer
    Pack size 100ml
    Form Solution (liquid)
    Applications Western blotting, ELISA, immunofluorescence, Immunoprecipitation; protein purification; SDS-PAGE; protein assays; epitope tagging; reporter gene analysis

    Broad Spectrum Protease Inhibitor Cocktail (EDTA free) (100x) (AR1182-1)

    • Specific References (4)
    SKU AR1182-1
    Pack size1mL
    Physical StateLiquid, colorless, transparent
    Description Boster’s Broad Spectrum Protease Inhibitor Cocktail (EDTA free) is a complex of various protease inhibitors, which has been tested for inhibiting proteases and esterase broadly. 

    Broad Spectrum Phosphatase Inhibitor Cocktail (AR1183)

    • Specific References (26)
    SKU AR1183
    Product nameBroad Spectrum Phosphatase Inhibitor Cocktail
    Pack size 1 mL
    Applications WB sample preparation, protein purification, Co-IP, protein kinase activity assay experiments

    Broad Spectrum Protease Inhibitor Cocktail (100x) (AR1182)

    • Specific References (44)
    • Q&As (2)
    SKU AR1182
    Pack size 1mLX2
    Form Solution (liquid)
    Applications Western blotting, protein purification, Co-IP; assays for protein expression, activity, modification, profiling and characterization, quantitative measurement; epitope tagging; reporter gene analysis

    RIPA Lysis Buffer (AR0105-100)

    • Specific References (73)
    • Q&As (2)
    SKU AR0105-100
    Product name RIPA Lysis Buffer
    Pack size 100ml
    Form Solution (liquid)
    Applications Western blotting, ELISA, immunofluorescence, Immunoprecipitation; protein purification; SDS-PAGE; protein assays; epitope tagging; reporter gene analysis

    Cytoplasmic and Nuclear Protein Extraction Kit (AR0106)

    • Specific References (40)
    • Q&As (4)
    SKU AR0106
    Pack size 1 kit
    FormLiquid

    Complete Bacterial Protein Extraction Reagent (AR0157)

    • Q&As (1)
    SKU AR0157-100
    Pack size 100mL (sufficient for lysis of 20g of bacterial biomass)
    Form Liquid

Displaying 7 Items