Whenever I can I like to dilute standards with the same buffer used for my sample to avoid matrix effect. Also, if you work frequently with one protein it would be useful to have a standard composed of the protein you want to measure rather than BSA; in this case accurate protein concentration of the standard should be determined by amino acid analysis.
Dear Heba,
Check this protocol attached below
During my PhD work I used PBS to dilute
I hope this will help
http://ergo.berkeley.edu/be115/BCA.pdf
The BSA standards can be diluted with water. For my BCA assays, I dilute the standards to final concentrations: (0, 250, 500, 1000, 1500 and 2000μg/m
L) to generate a standard curve and determine an absorbance- concentration relationship. Some researchers plot a linear regression which unfortunately is inaccurate. I prefer using the curvilinear regression to determine unknown protein concentrations of my samples instead.
This technical tip from Thermoscientific should give you better insight:
https://tools.thermofisher.com/content/sfs/brochures/TR0057-Read-std-curves.pdf
Whenever I can I like to dilute standards with the same buffer used for my sample to avoid matrix effect. Also, if you work frequently with one protein it would be useful to have a standard composed of the protein you want to measure rather than BSA; in this case accurate protein concentration of the standard should be determined by amino acid analysis.
BSA is hydrophobic, however it dissolved well in slightly luke warm distilled water or PBS (pH 7.2). Use odd series of concentration of BSA say 10, 30, 50, 70, 90 and 110 microgram/ml to calibrate a reference profile of BSA. Its always better to prepare BSA in the same buffer in which you have the actual/ test protein present.
- Badges
- Science topic
- Similar topics
- Chemistry
- Biochemistry