Column Capacity study [August 10, 2004]
Posted: Sun Aug 29, 2004 6:37 pm
By Apharmd Battler on Tuesday, August 10, 2004 - 09:40 am:
I'm new to method development and new to the world of protein chromatographic seperation. I've been asked to complete a column capacity study and a little lost as to how I am to go about this. Everyone area is swamped and assistance is minimal. I have a couple of columns they'd like for me to take a look at. I guess I need to know how should I go about getting started? Any suggestions would be greatly appreciated.
-------------------------------------------------------------------------------------------------------
By yiya on Tuesday, August 17, 2004 - 01:46 pm:
probably you need to provide us you column type and target analyte type.
-------------------------------------------------------------------------------------------------------
By A.Mouse on Tuesday, August 17, 2004 - 03:36 pm:
Also please explain wht you mean by column capacity study!
-------------------------------------------------------------------------------------------------------
By Chris Pohl on Saturday, August 21, 2004 - 05:35 pm:
There are generally two ways to characterize a protein separation column for loading capacity: measurement of breakthrough capacity and measurement of dynamic loading capacity. Do you know which one they are interested in having you measure? Breakthrough capacity is easiest to measure but has the least practical value since it greatly overstates the actual capacity under practical conditions. All that is required is that you pump a solution of known concentration through the column while connected to a UV detector at a suitable wavelength for detection of the protein of interest. The capacity is determined by measuring the breakthrough volume (the point at which the protein is found to exit the column). If you are interested in measuring dynamic capacity, this is generally done by increasing the injection concentration of a given protein until substantial peak broadening is observed. Generally, a 10% decrease in efficiency is identified as the loading limit in such a measurement.
I'm new to method development and new to the world of protein chromatographic seperation. I've been asked to complete a column capacity study and a little lost as to how I am to go about this. Everyone area is swamped and assistance is minimal. I have a couple of columns they'd like for me to take a look at. I guess I need to know how should I go about getting started? Any suggestions would be greatly appreciated.
-------------------------------------------------------------------------------------------------------
By yiya on Tuesday, August 17, 2004 - 01:46 pm:
probably you need to provide us you column type and target analyte type.
-------------------------------------------------------------------------------------------------------
By A.Mouse on Tuesday, August 17, 2004 - 03:36 pm:
Also please explain wht you mean by column capacity study!
-------------------------------------------------------------------------------------------------------
By Chris Pohl on Saturday, August 21, 2004 - 05:35 pm:
There are generally two ways to characterize a protein separation column for loading capacity: measurement of breakthrough capacity and measurement of dynamic loading capacity. Do you know which one they are interested in having you measure? Breakthrough capacity is easiest to measure but has the least practical value since it greatly overstates the actual capacity under practical conditions. All that is required is that you pump a solution of known concentration through the column while connected to a UV detector at a suitable wavelength for detection of the protein of interest. The capacity is determined by measuring the breakthrough volume (the point at which the protein is found to exit the column). If you are interested in measuring dynamic capacity, this is generally done by increasing the injection concentration of a given protein until substantial peak broadening is observed. Generally, a 10% decrease in efficiency is identified as the loading limit in such a measurement.