Protein Concentration assay (Bradford assay)
Posted 09 September 2009 - 11:46 AM
I got some problems by the protein concentration and I am not sure if I am doing it right:
I make the standard concentrations using BSA (0,5,10,20,40,60,80,100 ug/ml) and add 1 ml Bradford reagent. I take 1uL of my sample and add to 999uL water and 1ml Bradford. I take 300uL of standards and samples put them into 96well plate and read at 570nm.
then I draw the standard curve and find tthe concentration of my samples. Because I want at least 50ug protein for my further test (caspase3) I make this calculation:
(50ug*1uL)/Xug(obtained from standard curve)=
Is it all right? It seems that sth is missing. Do I have any dilution factor or sth to multiply?
Thank you for any advice
Posted 09 September 2009 - 02:42 PM
I'm horrible at calculating dilutions out by hand, but I can figure out the formulas to put into a spreadsheet! If you're looking for an explanation, try this tech sheet at Thermo.
Posted 09 September 2009 - 02:49 PM
C1*V1 = C2*V2, where C2 = concentration of the new solution.
C1 = (say) 32.7ug
V1 = 1uL
C2 = 50ug
V2 = ?
It's easy to figure out.
unless i've read the first post wrong.