Judith Sunzenauer (last update 1.4.2015)

Procedure
Preparation of Samples and Standards:

• Heat incubator to 37 °C
• Prepare plate layout with standard and sample dilutions in duplicates
• Thaw protein lysate samples
• Dilute protein lysate samples 1:10 (1:100) with RIPA buffer according to expected protein concentration

• Prepare dilutions of Albumin Standard Stock Solution (2mg/ml) with 1x PBS:
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Preparation of Working Reagent and Measuring:

• Mix 50 parts of BCA Reagent A with 1 part of BCA Reagent B (50:1). 200 µl of the Working Reagent (WR) is required for each well in the microplate.
• Pipette 10 µl of each unknown sample and standard into a flat bottom microplate
• Add 200 µl of the WR to each well and mix plate thoroughly on a plate shaker for 30 seconds
• Cover plate and incubate at 37 °C for 30 minutes
• Allow plate to cool to room temperature and measure the absorbance at or near 562nm on a plate reader. (Wavelengths form 540-590nm have been used successfully)

Material