Why should you use double referencing?

to correct for ligand versus reference differences
to correct for blank (buffer only) injections
to correct for bulk differences in the analyte
to correct for drift during the analyte injection

Why should you use the 'Kinetic / high performance' inject method?

to separate the sample from the flow buffer
to have slow injections
to have lower bulk shift jumps
to have better dissociation curves

With a slow dissociation, the best set up is to use

multi cycle kinetics
single cycle titration kinetics
short & long kinetics
steady state kinetics

Randomization of the samples is done because

it minimizes systematic errors
it can reveal systematic errors
it proves the system is reproducible
it minimizes carry over between samples

To get a better estimation of the kinetics you

repeat the method on the same sensor chip
use a wide analyte concentration range
add more analyte injections in the method
repeat the method with different ligand and analyte batches

How do you detect mass transport limitation?

by immobilizing less ligand
by varying the flow rate during analyte injection
by injection several analyte concentrations
by varying the analyte injection time

What can you try to minimize non-specific binding?

use a low pH running buffer
use a higher flow rate
lower the analyte concentration
adding more salt or detergent to the running buffer

What can you do when there is baseline drift?

use a higher flow rate
immobilize less ligand
flow for a longer period of time with flow buffer
ad more salt or detergent to the running buffer

What is the best analyte concentration range to inject?

0 – 1 µM
0 – 100% of Rmax
0.1 – 10 times the expected KD
at least one concentration should saturate the ligand

Replicate analyte injections are done

to prove the system is stable and reproducible
to have more curves to analyse
to reveal dilution errors
to minimize systematic errors

Why should replicates be in separate capped vials?

to minimize sample dilution
to minimize sample evaporation
to minimize carry over between samples
to minimize sample loss to the vial wall

Some SPR instruments use a calibration routine. Why?

to initialyse the fluidics
to get the best detector sensitivity
to make sure the curves are separated well
to make sure the system is equilibrated

Which of the statements is not true?

A slow dissociation needs a longer dissociation time than a fast dissociation to resolve the kinetics
To determine the Rmax, the ligand has to be fully saturated
SPR can be used for both high and low affinity interactions
Both the association and dissociation curve follow an exponential curve

Why should the buffers used be filtered and degassed?

To prevent air spikes and drift
To minimize the bulk effect
To have a better buffer equilibration
To have a better 1:1 interaction

To determine the k_a and k_d of an interaction you need at least

One analyte injection with 1x K_D concentration + dissociation
One analyte injection that is saturating the ligand + dissociation
Two analyte injections to steady state + dissociation
Two analyte injections with curvature + dissociation