Western Blot Protein Quantification Assays


Introduction

After protein sample preparation, the next step of western blotting is total protein analysis. Total protein analysis is the process of quantifying the protein concentration. Protein concentration data is critical to getting good results on your blot: equal amounts of protein must be loaded into each well of the gel to avoid saturation issues, ghost bands, and other trouble. The concentration data produced by total protein analysis can also perform as a reliable loading control for semi-quantitative western blotting. Boster provides a range of high-quality protein analysis kits so you can get more accurate protein concentration data and better blots.

Procedure

To quantify the protein using the BCA protein assay kit, the first step is to reconstitute the bovine serum albumin standard samples with 0.9% NaCl or PBS solution to create a solution with a known concentration. After preparing the BSA standard solution, create a gradient dilution using the BSA standard solution and the diluent solution included in the kit. Prepare the BCA working reagent using a 50:1 reagent A:B ratio and add it to the BSA gradient dilution. Incubate at room temperature and measure the absorbance of the samples at 562 nm. Plot the standard curve to determine the protein concentration of your protein sample.

For a detailed protocol for total protein analysis, see our Western blot handbook.

Tips

  1. Select the right protein assay for your application; choose between silver staining reagents, BCA assays, and commassie blue stains.
  2. Avoid inconsistent band strengths across wells, streaked bands, and saturation problems by getting accurate protein concentrations and optimizing loading amounts. See our troubleshooting guides for more information.


Displaying 5 Items

    Commassie Blue Staining And Destaining Solution (AR0163)

    SKU AR0163
    Pack size1kit (for 10 assays to stain gels of 5 x 8.5cm)
    FormLiquid
    ApplicationsFor protein staining on polyacrylamide gels.

    Silver Stain Kit (AR0171)

    • Specific References (1)
    • Q&As (2)
    SKU AR0171
    Pack size 1 kit (for 30 assays to stain the gel of 5 X8.5)
    FormLiquid
    Application To detect protein in polyacrylamide gels

    Micro BCA Protein Assay Kit (AR1110)

    • Specific References (17)
    • Q&As (2)
    SKU AR1110
    Pack size 1 kit
    Applications Western blotting, protein expression assays, protein profiling and characterization, protein quantitation assays

    BCA Protein Assay Kit (AR0146)

    • Specific References (262)
    • Q&As (1)
    Catalog numberUnit SizePrice(USD)
    AR0146-500Tube procedure: 250 assays
    Microplate procedure: 2500 assays
    108
    AR0146-1LTube procedure: 500 assays
    Microplate procedure: 5000 assays
    167

    Coomassie (Bradford) Protein Assay Kit (AR0145)

    • Specific References (7)
    • Q&As (1)
    SKU AR0145
    Product name Coomassie (Bradford) Protein Assay Kit(for 100-1500 μg/ml protein)
    Form Solution (liquid)
    Applications Western blotting, ELISA, immunofluorescence; protein expression assays, protein profiling and characterization; protein quantitation assays

Displaying 5 Items