KD increased observation

Hello, I am a little puzzled about the KD recovery of this Ab. About 3 months ago, I got 2.87E-10 M for KD. ka: 5.99E5, kd: 1.71E-4, Rmax: 129.7. Then about 3 months later, I got 4.20E-10 M for KD. ka: 4.29E5, kd: 1.80E-4, Rmax 129.6. I have been getting this KD, historically between 0.06 nM - 0.39nM. Now it is slightly out of range. Conc. of the Ab is at 4ug/mL. Nothing has changed between the 2 runs. Same chip, same buffer conditions, same Ab lot, same Ag lot. Any comments on why the difference? See attached sensorgrams. I would say though that the Ab is about 6 years old, kept frozen at -20C. Thank you.

So your new K_D value is 0.42 nM and you have already a range of 0.06 - 0.39 nM. What is the mean of the range and the standard error? Maybe you are too over concerned about this measurement.
And did you really use the same buffer after 3 months? When you make the same buffer fresh there are small differences in buffer composition and these will give small differences in the measurements. In my opinion the difference is too small to be worried about it.
Arnoud

Sorry for the very late reply. The buffer was >3 months old. Thanks for your input.