GE HEALTHCARE Biacore T100, Handbook

Page: 186 / 238

9 Kinetics and affinity analysis
9.8 Predefined models
186
Biacore T100 Software Handbook BR-1006-48 Edition AE
9.8.5 Kinetics – Two State Reaction
This model describes a 1:1 binding of analyte to immobilized ligand followed by
a conformational change that stabilizes the complex. To keep the model simple,
it is assumed that the conformationally changed complex can only dissociate
through the reverse of the conformational change:
A + B = AB = AB*
Note that conformational changes in ligand or complex do not normally give
a response in Biacore.
A good fit of experimental data to the two-state model
should be taken as an indication that conformational properties should be
investigated using other techniques (e.g. spectroscopy or NMR), rather than
direct evidence that a conformational change is taking place.
Rmax2 Analyte binding capacity of the second ligand
(RU) Rmax2
Conc Analyte concentration (M) Conc
tc Flow rate-independent component of the
mass transfer constant tc
Flow Flow rate (µl/min) f
kt Mass transfer constant tc
×
f
1/3
RI Bulk refractive index contribution in the
sample RI
Model parameters Obtained from
ka1 Association rate constant for analyte binding
(M
-1
s
-1
)
Fitted
kd1 Dissociation rate constant for analyte from
the complex (s
-1
)
Fitted
ka2 Forward rate constant for the conformational
change (s
-1
)
Fitted
kd2 Reverse rate constant for the conformational
change (s
-1
)
Fitted
Rmax Analyte binding capacity of the surface (RU) Fitted
Conc Analyte concentration (M) Provided as
input
tc Flow rate-independent component of the
mass transfer constant Fitted