Application Guide

October 1, 2022
Keywords or phrases:
Quality control, GxP biotherapeutics,
process development, product attributes,
design of experiments

Quality Control of Biotherapeutics
Using Octet® Systems
David Apiyo, Sartorius, Fremont, CA
Correspondence
Email: octet@sartorius.com

Abstract
The Octet® family of Bio-Layer Interferometry (BLI) systems are used throughout biotherapeutic discovery, development,
manufacturing, and quality control (QC) workflows to simplify and streamline measurement of process and productattributes. These systems offer an excellent alternative to assays performed using time- and labor-intensive methods suchas ELISA and HPLC. Octet® assays are label-free, fully automated, require much less user intervention, and provide asimplified workflow.
All Octet® systems are GxP ready and are widely adopted in regulated environments. The microplate-based sample format
is ideal for Design of Experiments (DoE), and, ready-to-use or customizable biosensors create a robust and flexible platformthat can also accommodate direct analysis of crude samples.

Find out more: www.sartorius.com

Bio-Layer Interferometry (BLI) has been rapidly adopted as
an important analytical tool in laboratories that work withbiological molecules, either as drug products, vaccines, ordiagnostic reagents. Widely used and accepted in research
and assay development, BLI assays are now also utilized for
quality control (QC) in biologics development workflows forligand binding and product quantitation during lot releaseand in-process testing.
Octet® systems combine real-time binding analysis with asimple Dip and Read approach for rapid assay and method
development. For example, the 8-channel Octet® R8system quantitates 96 mAb samples in only 30 minutes,compared to >6 hours by ELISA1 or as long as 10 hours via
HPLC, resulting in significant project cost and time savings.
Optimizing a generic potency assay takes approximately
half the time on Octet® systems as it would with surface
plasmon resonance (SPR) or on manual ELISA.
Octet® systems also offer several other advantages overcomparative analytical technologies. The microplate-based
sample format is ideal for Design of Experiments (DoE),
enabling rapid, high-throughput method optimization.
Disposable, ready-to-use or customizable biosensors are
simply dipped into samples for analysis – no priming
needed. This lack of microfluidics results in a more robust
and flexible platform that can also accommodate direct
analysis of crude samples.

Is the Octet® Platform GxP Ready?

---All Octet® instrument models are GxP ready. Sartorius
offers GxP users an assortment of products and services
that ensure full confidence in data integrity. They include:
21 CFR Part 11-compliant software that can be used on allOctet® models including Octet® R series, RH16 and RH96
Instrument Installation and Operational Qualification(IQ/OQ) Kits
Performance Qualification (PQ) Kit validated for theOctet® R8 instrument
High Precision Streptavidin 2.0 Biosensors (SAX2) qualified for seamless use across multiple lots of biosensors
Support service for customer-run software validation

Is the Octet® Recommended for
Use in Regulated Environments?
Regulatory bodies such as the FDA, EMEA, NMPA andothers do not typically recommend or refer to specificproduct, brand, or vendors in their guidance on analyticaltechnologies. While there may be references to certain
technologies in some USP chapters, these are references2

based on general concepts around the application inquestion. Label-free methods such as BLI and SPR are
simplified immunoassays based on the ELISA-type binding
assay principle, so their use for measurement of potency or
product quantity is based on decades of established andaccepted measurement principles. Binding kinetics
constant measurements using label-free methods arealso well established, and BLI technology has been
demonstrated through use in thousands of laboratories
world-wide and in peer-reviewed publications. In fact, bestpractice in the drug development industry employs
orthogonal or alternative platforms that are fit for purpose.
The key factor here is that the sponsor sets appropriate
acceptance criteria and that the technology in use enables
the development of suitable methods that are robust andmeet the criteria for quality control for the specific product.

Which Octet® Biosensors are Suitablefor QC Methods?
The selection of biosensor surface chemistry depends on
the application. All Octet® Biosensors can be used to develop QC methods. However, during assay method
development, repeatability, intermediate precision andreproducibility studies that include biosensor lot to lot
assessment should be done to determine assay robustness
in line with recommendations from the relevant regulatory
bodies. High Precision Streptavidin 2.0 Biosensors (SAX2)(Figure 1), intended for use with biotinylated ligands, havebeen developed to ensure minimal lot to lot variations inligand capture. This biosensor is recommended for use withany assay where high ligand immobilization reproducibility
is critical. The SAX2 Biosensor is suitable for both ligand
binding kinetics assays and for custom quantitation assays(through pre-coating of a capture ligand on the biosensor).

What If I Have a Different Octet® Modelin Development Than the One in QC –Is Octet® Data Comparable?
Analytical method changes are a normal process in theproduct life cycle and can be triggered by a variety offactors such as the need to better understand productquality or to replace legacy methods, and other reasons. In
any case, regulatory bodies require that bridging studies bedone when a method change is necessitated and that thenew method be validated. Although bridging studies are arequirement, the data obtained on any Octet® system –from the Octet® RH96 to the RH16 or the 8-channelOctet® R8 – are comparable so long as the acquisition
parameters are of high precision.

A.

Loading

AssociationB.

Dissociation

2.0

Global ﬁtting for kinetics

1.0

1.6

0.8
Binding (nm)Binding (nm)

1.2

0.8

0.6

0.4
0.4
0.2
000

400

800

1200

1600
Time (sec)

2000

2400

02800

200

400

600

800
1000
Time (sec)

1200

1400

1600

1800

Figure 1: (A) Kinetic analysis of the interaction between a ligand biotin-Fcγ Receptor (~55 kDa) and analyte hIgG (150 kDa) with SAX2 Biosensor,
overlaying three lots of SAX2 and raw data aligned at the ligand loading step. (B) Data was processed and curve fitted using a 1:1 binding model.
See the Sartorius datasheet, High Precision Streptavidin 2.0 (SAX2) Biosensors for ligand loading specifications.

Figure 2 and Tables 1 and 2 show exemplary kinetics
characterization data for PSA antigen binding to its
antibody as derived from the three different Octet® models.
The data suggests minimal differences in response signals
for both ligand capture and antigen binding, resulting in
5A.

good precision in the extracted affinity constants. Thisdata demonstrates that Octet® systems deliver consistent
performance across the platform, regardless if researchers
are using multiple, same-model instruments, or different
Octet® instruments.

B.

0.6

4Binding (nm)Binding (nm)

0.4
320.2

1000C.

20

40

60

80
100
Time (sec)

120

140

160

D.

0.6

0200

400

600
Time (sec)

800

1000

1200

0200

400

600
Time (sec)

800

1000

1,200

0.6

0.4
Binding (nm)Binding (nm)0.40.20.2000200400600
Time (sec)

8001000

1200

Figure 2: Octet® comparability kinetics studies showing side by side kinetics data for biotinylated anti-PSA antibody immobilized on two different
lots of SAX2 Biosensors binding to PSA antigen. (A) An overlay of the immobilization data from three instruments where PSA antigen was used at
200 nM, 100 nM, 50 nM, 25 nM and 12.5 nM using the Octet® R8 (B), Octet® RH16 (C) and Octet® RH96 (D) systems. Data was acquired in duplicates for each biosensor lot. Only one lot was used on the Octet® HTX system. Experiments were run at 25°C using the 8-channel read head mode
for all three Octet® models.
3

Table 1: Comparison of biotinylated anti-PSA ligand response on different
Octet® models. Data was obtained from two lots of SAX2 Biosensors in
duplicate (see Figure 2A). Loading response is reported at 150 seconds.
Octet® model

Mean response(nm)

Range, nm(high-low)% CV

R81.501.59–1.42

3.16

RH16

1.67

1.73–1.61

2.04

RH96

1.68

1.74 –1.641.78

Reference
1.

Biolayer Interferometry as an Alternative to HPLC for
Measuring Product Concentration in Fermentation
Broth, Rathore AS et al., LCGC, 35(12) 870–877.

Table 2: Octet® kinetics comparability study data shows kinetics parameters for biotinylated anti-PSA antibody immobilized to two different lots of
SAX2 Biosensors binding to PSA antigen. Characterization was performedin duplicate for each biosensor lot and on all three Octet® instruments
using the 8-channel read head. Experiments were run at 25°C.
Octet® modelBiosensor
lot 1 KD (M)Biosensor
lot 2; KD (M)Mean,KD (M)

% CV

R8

6.91E-09

5.79E-09

6.35E-09

12.5

RH16

7.46E-09

7.13E-09

7.29E-09

3.25

RH96

7.71E-09

n.d.

7.71E-09

n.d.

Inter-instruments(mean)

7.36E-09

6.4565E-09

7.12E-09

9.79

Germany
Sartorius Lab Instruments GmbH & Co. KG
Otto-Brenner-Strasse 2037079 Goettingen
Phone +49 551 308 0F
 or 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.
4006 Rev B

USASartorius Corporation565 Johnson Avenue
Bohemia, NY 11716
Phone +1 888 OCTET 75
Or +1 650 322 1360