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
SKU AR0101
Pack size 100ml
Form Solution (liquid)
Applications Immunoassays: WB, ELISA, RIA, immunoprecipitation; reporter assays; protein kinase assays; protein purifications; folding studies; chromatographic studies; DNA-protein interaction assays

Availability: Ships in 5-7 business days

SKU AR0056
Pack size 5 mL
Form Solution (liquid)
Applications Proteolytic induced epitope retrieval (PIER); Proteolytic inactivation of endonucleases during DNA/RNA isolation and tissue sections preparation for in-situ hybridization; Determination of enzyme localization; General protein digestion; Protein modification; Improve cloning efficiency of PCR products
Availability: Ships next business day
SKU AR1183
Pack size 1 ml, 100X concentrated stock solution
Description:Broad Spectrum Phosphatase Inhibitor Cocktail inhibits most phosphatases activities (details in datasheet).
Applications WB sample preparation, protein purification, Co-IP, protein kinase activity assay experiments
Availability: Ships next business day
SKU AR1182
Pack size 1 mL
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
Availability: Ships next business day
SKU AR0105
Pack size 50ml
Form Solution (liquid)
Applications Western blotting, ELISA, immunofluorescence, Immunoprecipitation; protein purification; SDS-PAGE; protein assays; epitope tagging; reporter gene analysis
Availability: Ships next business day
SKU: AR1121
Pack size:80mg, enough for 100ml of samples
Description:Phosphatase Inhibitor Cocktail III inhibits Serine and threonine phosphatases; acid and alkaline phosphatases; PP1 and PP2A activities (details in datasheet).
Applications: WB sample preparation, protein purification, Co-IP, protein kinase activity assay experiments
Availability: Ships next business day
SKU:AR1120
Size:80mg, enough for 100ml of samples
Description:Phosphatase Inhibitor Cocktail II inhibits Tyrosine protein phosphatases, acid and alkaline phosphatases. It is used to protect samples against endogenous phosphatase activies so that downstream assays can accurately study the phosphorylation profile of the sample.
Application:WB sample preparation, phosphorylation studies
Availability: Ships next business day
SKU AR0156
Pack size 1 kit, sufficient for 50 assays
Applications Mitochondria Isolation
Description Mitochondria Isolation Kit, Used to isolate the intact and purified mitochondria from animal cells and tissues, and prepare the mitochondria protein of animal tissues and cultured cells.
Availability: Ships next business day
SKU AR0155
Pack size 1 kit, enough reagents for 50-100 assays
Kit Components Protein Extraction Solution A 30ml
Protein Extraction Solution B 30ml
Applications Immunoassays: WB, ELISA, RIA, immunoprecipitation; reporter assays; protein kinase assays; protein purifications; folding studies; chromatographic studies; DNA-protein interaction assays
Availability: Ships next business day
SKU AR0106
Pack size 1 kit, enough reagents for 50-100 assays
Kit Components Cytoplasmic Extraction Reagent (CER) 30ml
Nuclear Extraction Reagent (NER) 5ml
Applications Immunoassays: WB, ELISA, RIA, immunoprecipitation; reporter assays; protein kinase assays; protein purifications; folding studies; chromatographic studies; DNA-protein interaction assays
Availability: Ships next business day
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