can you please explain how to prepare x and how to calculate the initial concentration of x in your exemple because i didnot realy get the calculation from the dilution
Shouldnt the curve be starting from the beginning of the two axes (point 0,0)? Because if you have no absorbance you cant have concentration. And also, isnt the standard curve a version of Lambert-Beer's law? If so, we cant have equation of y=ax + b formation, we will have the formation y=ax. And the thing i cannot understand is why we do the standard curve. Cant we just use the bradford reagent and measure the complex absorbance and after that, using Lambert-Beer's law we find the protein's concentration? Please correct me if im wrong.
As far as I understand for the Lambert-Beer's equation (A=C*e*l) you would need to use the extinction coefficient which is different for each reagent and is determined at a specific wavelength (defined by the solution you are measuring). If you observe, here they are using a 96 well plate here and is being read by the luminometer, I guess this is the instrument they will use to determine the concentration of proteins for their samples, but it can be done with a UV-vis. For the A=C*e*l normally you use an UV-vis spectrophotometer with a 1 cm cuvette, which allows you to set l=1 cm in the LB equation. (Or if you use a different pathway length I guess you would need to take that into consideration, for instance, if you are going to measure the concentration with a nanodrop, which you would need to adjust the value for "l", but anyway, I would strongly advise you to always use a cuvette to have more reliable results.
Cause u can plot the linear data and get an ecuation from that. Then, u use that ecuation to determine the concentration of your protein. If u use non-linear data, would be harder to use a good plot ecuation
