I have BHQ-COOH and I would like to bind it to a certain protein. I want to calculate the exact amount of BHQ I would need to add to the protein so I dont have to filtrate every time. I cannot do molar ratios obtaining the absorbance values since I dont know the stichiometry. Does anyone know how to approach this?
You can measure the concentration of a dye solution by its absorbance using Beer's law if you can find out the extinction coefficient for your particular dye in the solvent you have it in.
This method can also be used to measure the dye concentration in a protein solution once the dye has been conjugated to the protein and any excess dye has been removed. Combine this dye concentration measurement with a protein concentration measurement (such as by BCA, Bradford, or Lowry method), and you can calculate the labeling stoichiometry.
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