Pierce™ BCA Protein Assay Protocol
Materials and Equipment:
- Pierce™ BCA Protein Assay Kit (A55864)
- Sample (protein sample)
- BSA (Bovine Serum Albumin for creating a standard curve)
- Microplate (96 wells)
- Multi-pipette (for dispensing samples and standards)
- Spectrophotometer or plate reader
Steps:
1. Add your protein samples to each well of the microplate. Add 25-200 μg of protein to each sample well.
2. Dilute the standard solution to various concentrations, typically 0, 7.8, 15.6, 31.25, 62.5, 125, 250, 500 μg/mL, etc.
3. Prepare the Pierce™ BCA Reagent mixture: Mix 50 parts of Pierce™ BCA Reagent Solution A with 1 part of Pierce™ BCA Reagent Solution B. For example, mix 50 μL of Solution A with 1 μL of Solution B to create the working Pierce™ BCA Reagent solution. (A:B=50:1)
4. For each standard concentration and sample, add the Pierce™ BCA Reagent and mix well. Incubate the plate at 37°C for 30 minutes.
5. After the incubation, measure the absorbance at 562 nm using a spectrophotometer or plate reader.
6. Create a standard curve using the absorbance values for each standard. Plot the data and perform linear regression analysis.
7. Calculate the protein concentrations of your samples using the standard curve and the absorbance values obtained.
