Hello,What is your difference between the maximum and minimum value ? Have you try to change the probe concentration?Best,DidierLe 21 juil. 2017 15:34, "Mohammad Khan" <[log in to unmask]> a écrit :Dear all,I am trying to measure the difference in polarization upon the binding of the DNA to my protein. I take 1-5 nM of Cy3-labelled DNA and add varying dilutions of my protein to it (100 microM to 1 nM). I do get a decrease in difference of polarization with decrease in protein concentration. However, the results are difficult to reproduce and also vary greatly within triplicates of an experiment.Similar observations have been observed by my colleagues with their proteins.Are there any tips or precautions to keep in mind while setting up these reactions?Looking forward for suggestions.Thank you.