Bio-Rad Foresight™ Chromatography Columns, Prepacked - Manual (Page 13)

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Note:
1. The slope of the operating line is equal to the phase
ratio ( β).
2. The feed concentration and the phase ratio determine
whether, after enough incubation time (i.e., 4–6 hours),
media saturation will be attained (plateau of the
isotherm).
3. A small volume of media relative to a fixed sample
volume is recommended for capacity studies.
4. A larger volume of media relative to a fixed sample
volume is recommended for wash and elution studies.
In sum, it is important to bear in mind that the capacity
utilization for a given chromatographic medium is a
function of the shape of the underlying solute isotherm
and the magnitude of the phase ratio.
Several models are available to model an isotherm. 10–11
A model that is frequently used to represent adsorption
isotherm data is the single component Langmuir isotherm
Eq. 7.
1 m
Kc
q q
Kc
=
+
Where q
m
represents the maximum binding capacity, K
represents the association constant; q and c represent the
adsorbed and liquid phase concentrations at equilibrium.
The two model parameters (q
m
, K) can be estimated by
parameter regression, for example in Excel using the
solver add-in tool.
2.5 Uptake Curve
The uptake curve is constructed by taking time
points along the operating line.
These points are created by loading the sample in
different wells at predetermined time intervals.
The sample to be incubated the longest is added
first.
Eq. 7