High U value in some runs

Hi
what is the reason for very high U value (around 95) only in some runs/samples. Could someone please help me in understanding the concept of U value in detail as some of my runs/samples are giving high value in the developed method while some are giving low values for U value (within limits of 25).
What is its relation to association and dissociation??
Why it fails only in some runs??

I am running Mabs as analyte with Fcgr3aV158 as the receptor. Fcgr3a is captured on the anti his immobilised chip. Capture levels are fairly constant throughout the runs.

Hi,

The parameter uniqueness value or U-value is a value that reflects the correlation between k_a, k_d and Rmax.

In some situations, it may be possible to determine a value for a combined function of two or more parameters without being able to determine unique values for the individual parameters. Such parameters are said to be correlated.
One example is the pair of kinetic rate constants k_a and k_d, that are related through the affinity constant K_D: it may be possible to determine the affinity constant reliably without being able to resolve the individual rate constants.
Parameter uniqueness is assessed by testing correlation between pairs of the parameters k_a, k_d and Rmax. If significant correlation is found, this will be reported as a warning that parameters cannot be uniquely determined.

Note: This test does not explore all possible parameter correlations. A Pass status for this test is not a fail-safe indication that parameters are uniquely determined.

Thus when the U-value fails, check the calculated values if they make sense. A new fit with other starting values may give a better fit and better values and a good U-value.
As stated above, the U-value is a tool to assess the uniqeness of the fit values it is not a validiation.

Kind regards
Arnoud

Hi Arnoud
As you suggested, I have checked other parameters also. For the samples that have passed for U value, ka, kd and kD makes sense and the Rmax is also fine. But for the samples for which U value is high, Rmax is very low and ka, kd are also not giving proper values. Binding is less in these samples. Could this be a sample issue?? How do I check this??

Without seeing the two types of curves it is difficult to give usefull advice. Is the observed Rmax very low or does the fit report a low Rmax? When the association part has not enough curvature or the dissociation it too short the kinetic parameters cannot be uniquely determined.
Regards
Arnoud