Preparation of protein standards for BCA Pierce Assay: Tracey's method
Protein homogenates were in lysis buffer containing 1% SDS.
This is diluted: 10 ml + 40 ml water for protein assays. Therefore final concentration 0.2% SDS.
BSA standards come in 1 ml aliquots at 2000 mg/ml (2 mg/ml) (no SDS).
It is important that the standard is diluted such that the final concentration of SDS is equal to that in the samples to be tested i.e. 0.2% SDS.
TO DO THIS:
- Empty an aliquot of 2000 mg/ml standard into a 1.5 ml tube. Tube 1 = will be 1000 mg/ml stock.
- Remove 0.5 ml from tube 1 into a 2nd tube. Add 0.5 ml water. Tube 2 = will be 500 mg/ml stock.
- Mix tube 2 well and remove 0.5 ml into a 3rd tube. Add 0.5 ml water. Tube 3 = will be 250 mg/ml stock.
- Mix tube 3 well and remove 0.5 ml into a 4tjj tube. Add 0.5 ml water. Tube 4 = will be 125 mg/ml stock.
- Mix tube 4 well and remove 0.5 ml into a 5th tube. Add 0.5 ml water. Tube 5 = will be 62.5 mg/ml stock.
- Add 0.5 ml of 0.4% SDS in water to all 5 tubes. This brings all standards to final concentration of 0.2% SDS.
Use equal volumes of the standards and the samples.