Best Practice Guide

March 31, 2023
Keywords or phrases:
sensor chip hydration, normalization, priming,
conditioning

Best Practice Guide:
Octet SPR Sensor Chip Preparation®

Stuart Knowling Ph.D. Senior Scientist
Sartorius Corporation, 47661 Fremont Blvd, Fremont, CA 94538 USA
Correspondence
Email: octet@sartorius.com

Introduction
Octet® SPR sensor chips allow you to generate high-quality surface plasmon resonance (SPR) data and this requires that astable baseline is achieved prior to performing assays. An important parameter contributing to a stable baseline is a stablesensor chip surface. Best practice for preparing Octet® SPR sensor chips includes four basic steps that should be performed
sequentially prior to starting assays:
Normalization
Priming
Conditioning
Hydration
Octet® SPR sensor chips are supplied in a sealed packet containing an inert gas and should be stored at the recommended
condition upon delivery. Prior to installation, move the sensor chip to room temperature and allow to equilibrate without
opening the packet (~10 mins). Once temperature equilibrated the sensor chip is ready for installation.

Find out more: www.sartorius.com

Normalization

Priming

Normalization is an important step in sensor chip
preparation as individual sensor chips contain slightdifferences in their surface properties. Therefore, it isimportant to adjust the system’s detector to compensate
for these differences.

Priming is an important part of sensor chip installation as it
introduces running buffer to the flow cells of the sensor
chip and aids in rehydrating the sensor chip surface after
storage.

Normalization of the sensor chip should generally be
performed after the following events:
Installation of a new sensor chip for the first time
Installation of a sensor chip that has been previously
installed
When the assay temperature has been changed
The Octet® SF3 allows rapid normalization of the sensor
chip using 100% DMSO and is located under Instrument
Setup: Normalize.
Note: 100% DMSO: is typically used for normalization
on a new sensor chip which has not been previously
used or had a molecule immobilized on its surface. 70%
glycerol is typically used when normalization of sensor
chip with an immobilized ligand is required, this includessensor chips that contain pre-immobilized ligands suchas SADH.
Place the required solution in a sample rack as shown
below and drag across to the empty editor box:

The prime function is located under Instrument Setup:
Prime.
Best practice for priming is to perform a single prime whena new sensor chip is installed and normalized, and theconditioning steps shown below used. Three primes
should be performed when the running buffer is changed
to ensure the Octet® SF3 system and sensor chip areprimed into the new buffer.

Conditioning
Prior to immobilization of a ligand onto the sensor chip it isrecommended that the sensor chip is initially cleaned andhydrated to ensure the best results. Therefore, after priming
it is recommended that sensor chips are preconditioned
using the Fast injection type option with the following
conditioning solutions prior to immobilization:
COOH1, CDL, CDH and PCH

Item

Description

Position

Volume (µL)

Injection

SolutionContact
time (sec)

Flow rate(µL min-1)

Normalize

100% DMSO

R1A1

700

110 mM HCl

15100

250mM NaOH

15100

30.1% (w/v) SDS

15100

410 mM HCl15100550mM NaOH

15100

60.1% (w/v) SDS

15100Injection

SolutionContacttime (sec)

Flow rate(µL min-1)

11 M NaCl 50mM NaOH

15100

21 M NaCl 50mM NaOH

15100

31 M NaCl 50mM NaOH

15100

41 M NaCl 50mM NaOH

15100

70% Glycerol Normalization

SADH

When performing normalization on a sensor chip that has
material immobilized on it, 70% glycerol is recommended.
Item

Description

Position

Volume (µL)

Normalize

70% glycerol

R1A1700

It is important that the High Viscosity box is ticked whenusing 70% glycerol.

2HisCap
Injection

SolutionContacttime (sec)

Flow rate(µL min-1)

1400 mM EDTA

15100

2400 mM EDTA

15100

3400 mM EDTA

15100

4400 mM EDTA

15100

Following conditioning, it is best practice to perform asingle prime with the running buffer to help ensure astable baseline prior to performing further surface
preparation.

Hydration and Post-Conditioning

For example, in a multi-cycle kinetics assay with thefollowing parameters:
Association(Sec)Dissociation Flow rate(Sec)(µL min-1)

Analyte 0.411 – 100 nM

18060050Regeneration 100 mM HCl30NA50

Post-conditioning would consist of ~5 cycles:
Association(Sec)Dissociation Flow rate(Sec)(µL min-1)

Analyte 0.41 nM

18060050Regeneration 100 mM HCl

30NA50After immobilization, it is vital to ensure that temperaturefluctuations and more commonly, swelling of the sensor
chip matrix does not occur. This ensures that a stablebaseline is achieved prior to performing an SPR assay,
which leads to higher quality data. With the Octet® SPR
Discovery software this is easily achieved by including
startup cycles in methods.

Where OneStep® injections are performed, a 200-folddilution of the single concentration of analyte should beused.

Located in the Periodic Cycles section of the Method
Editor; startup cycles are performed at the start of the assay
and are not included in subsequent blank cycles.

With proper storage and preparation, Octet® SPR sensor
chips can be used to generate real-time kinetics andaffinity data for a wide range of molecules.

It is recommended that 5 – 10 startup cycles are includedin the method to help ensure a stable baseline prior toperforming the assay.

Conclusion

When combined with the best practices ofNormalization, Priming, Conditioning and Hydration,
which help ensure a stable sensor chip and baseline prior
to performing your method, Octet® SPR Sensor Chips
and the Octet® SF3 can generate high quality data foreven the most challenging assays.

It is recommended that post-conditioning of the sensor
chip is also performed prior to collecting data in order toremove any material that is not covalently attached to thesensor chip surface and to expose the sensor chip matrix
to potential pH shock prior to collecting data.
Post-conditioning is achieved by performing ~5 cycles ofanalyte binding (Association) and regeneration using thesame parameters as in the main assay. Post-conditioning
uses the lowest concentration of analyte and the same
regeneration solution to be assessed in the assay.

3Germany
Sartorius Lab Instruments GmbH & Co. KG
Otto-Brenner-Strasse 2037079 Goettingen
Phone +49 551 308 0

USA
Sartorius Corporation565 Johnson Avenue
Bohemia, NY 11716
USA
Phone +1 888 OCTET 75or +1 650 322 1360

For further contacts, visit
www.sartorius.com/octet-support

Specifications subject to change without notice.
Copyright Sartorius Lab Instruments GmbH & Co. KG.
For Research Use Only.
Octet-SF3-Sensor-Chip-Preparation-Best-Practice-Guide-en-L-Sartorius 4073 Rev B.
Status: 03 | 2023