Interaction only at slow flow rate ? - Forum


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TOPIC: Interaction only at slow flow rate ?

Interaction only at slow flow rate ? 18 May 2005 02:00 #1

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What does it mean if an analyte shows interaction with the ligand
only with a small flow rate, while it doesn't show interaction at higher
flow rate?
Ilaria

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Interaction only at slow flow rate ? 18 May 2005 02:00 #2

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what are the analyte and the ligand? what MW? protein or something else? which flow rates are used?

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Interaction only at slow flow rate ? 18 May 2005 02:00 #3

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my ligand is fibrin (MW: 340000) and the analytes are peptides with MW under 1000. I observed interaction at 5 microL/min but not at 30 microL/min. Why?

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Interaction only at slow flow rate ? 18 May 2005 02:00 #4

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Is the signal you get in correlation to the concentration saturable? Do you use a buffer containing Sodium chloride and polysobate as in biacore's HBS-EP buffer to limit unspecific binding? The ligand is quite small so I don't know how relevant transport limitation is. Another effect can be the influence of shearing forces on the protein (high shearing = conformational change that inactivates the protein). What happens if you measure at 10 and 20 ul/min. Is there some kind of flow-barrier where you loose all binding activity?
If you get nice results at flowrates around 10 - 20 ul/min I would go for this rates.

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Interaction only at slow flow rate ? 18 May 2005 02:00 #5

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How long did you inject your sample and how much RU change? Try to carefully maintain your Biacore system and degas the buffer and repeat the experiment again.

Try to increase the immobilized ligand density to see what happen.

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