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Bulk contribution messages
- OldForum
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13 years 3 weeks ago #1
by OldForum
Bulk contribution messages was created by OldForum
Hello....I've been having a lot of "High bulk contribution (RI) found" in the QC section of the Evaluation software. I tried setting the RI constant to zero, but that did not help the fit. Does anyone know what maybe going on? Where would this high bulk interference come from?
mtarca
mtarca
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- OldForum
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13 years 3 weeks ago #2
by OldForum
Replied by OldForum on topic Bulk contribution messages
I assume you use the new Biacore types and software (T100/T200).
I don't have experience with them but in general, bulk RI compensation in the fitting should be low.
First design the experiment, so that the bulk in the sensorgram is minimized.
The try this:
1) Try fitting the kd first.
2) Then fit the ka with the kd fixed (enter value from the first fit).
3) Then fit ka and kd (enter values from the first and second fit). Set RI to zero and don't fit.
Now you have good estimates of the ka, kd and the Rmax.
4) Try to fit floating ka, kd, Rmax and RI. Use the values from 3) as a starting point.
This stepwise fitting works the best with me.
High bulk compensation in the fitting is a sign that the fitting is not following the actual data. It is possible that the fitting can not find the true values for the parameters but it is also possible the data is not 1:1. First check for mass-transport, or lower the ligand concentration.
Maybe you can show a sensorgram with fitting.
Regards
Arnoud
I don't have experience with them but in general, bulk RI compensation in the fitting should be low.
First design the experiment, so that the bulk in the sensorgram is minimized.
The try this:
1) Try fitting the kd first.
2) Then fit the ka with the kd fixed (enter value from the first fit).
3) Then fit ka and kd (enter values from the first and second fit). Set RI to zero and don't fit.
Now you have good estimates of the ka, kd and the Rmax.
4) Try to fit floating ka, kd, Rmax and RI. Use the values from 3) as a starting point.
This stepwise fitting works the best with me.
High bulk compensation in the fitting is a sign that the fitting is not following the actual data. It is possible that the fitting can not find the true values for the parameters but it is also possible the data is not 1:1. First check for mass-transport, or lower the ligand concentration.
Maybe you can show a sensorgram with fitting.
Regards
Arnoud
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