GE HEALTHCARE Biacore T100, Handbook

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Biacore T100 Software Handbook BR-1006-48 Edition AE
181
Kinetics and affinity analysis 9
9.8.2 Kinetics – Bivalent Analyte
This model describes the binding of a bivalent analyte to immobilized ligand,
where one analyte molecule can bind to one or two ligand molecules. The two
analyte sites are assumed to be equivalent. The model may be relevant to
studies among others with signaling molecules binding to immobilized cell
surface receptors (where dimerization of the receptor is common) and to studies
using intact antibodies binding to immobilized antigen. As a result of binding of
one analyte molecule to two ligand sites, the overall binding is strengthened
compared with 1:1 binding. This effect is often referred to as avidity.
A +B = AB
AB + B = AB
2
Note: Once analyte is attached to the ligand through binding at the first site,
interaction at the second site does not contribute to the SPR response . For
this reason, the association rate constant for the second interaction is
reported in units of RU
-1
s
-1
, and can only be obtained in M
-1
s
-1
if a
conversion factor between RU and M is available. Similarly, a value for the
overall affinity or avidity constant is not reported.
Model parameters Obtained from
ka1 Association rate constant for the first site (M
-1
s
-1
) Fitted
kd1 Dissociation rate constant for the first site (s
-1
) Fitted
ka2 Association rate constant for the second site
(RU
-1
s
-1
)
Fitted
kd2 Dissociation rate constant for the second site (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
f Flow rate (µl/min) Provided as
input
tOn Sample injection start time (s) Provided as
input
tOff Sample injection end time (s) Provided as
input
RI Bulk refractive index contribution in the sample Fitted