Instructions For Use
USD 2937(c)
ScreenExpert RoboColumnu
For High Throughput Chromatography Sorbent Screening
and Optimization
Table of Contents
1.
Introduction ............................................................................................................................................ 3
2.
Characteristics ...................................................................................................................................... 3
3.
Applications ........................................................................................................................................... 4
4.
Advice on Handling ............................................................................................................................... 4
4.1 Equipment ....................................................................................................................................... 4
4.2 Sample Preparation ......................................................................................................................... 4
4.3 Experimental Set Up......................................................................................................................... 4
4.4 Te-Chrom Wizard ............................................................................................................................ 4
2
5.
Protocol
............................................................................................................................................... 6
6.
Column Scale-Up ..................................................................................................................................10
7.
Troubleshooting Guide ..........................................................................................................................10
8.
Ordering Information .............................................................................................................................11
1. Introduction
ScreenExpert RoboColumns are miniaturized chromatography columns prepacked with Pall chromatography sorbents. They are designed for fully automated and parallel chromatographic separations with
robotic liquid handling workstation such as Freedom EVOu workstation (Tecan) or Janus BioTx Pro Plus
workstation (Perkin Elmer).
ScreenExpert RoboColumns are packed with anion and cation exchange, mixed-mode or any other Pall
chromatography sorbents upon request. The columns are packed by Atoll GmbH and are identical to Atoll
MediaScoutu RoboColumns.
Bed containment between two filter frits and professional packing with consideration to individual material
compression requirements ensure high efficiency and peak symmetry similar to that of preparative and
process separation columns.
Up to 96 ScreenExpert RoboColumns can be individually and easily arranged onto a 96-well array plate.
As a result of this modular design, ScreenExpert RoboColumns technology is used in a broad spectrum
of applications, such as parallel screening and optimization of chromatographic conditions, scale-down
experiments, sample preparation.
ScreenExpert RoboColumns are supplied in a package of eight prepacked units with two removable
silicon cover seals for proper storage. A 96-well array plate is available to arrange up to 96 ScreenExpert
RoboColumn units.
ScreenExpert RoboColumns are ready-to-use 96-well formatted chromatography columns prepacked with
Pall chromatography sorbents. They are available in 200 μL and 600 μL sizes. Characteristics and available
column geometries are detailed in Table 1.
The chromatography sorbents used in ScreenExpert RoboColumns are also available in Pall PRC prepacked
columns of 1 mL and 5 mL, for the performance of scale-up studies.
2. Characteristics
Table 1
ScreenExpert RoboColumns Characteristics
ScreenExpert RoboColumns
Column volume
Bed height
Column inner diameter
Column material
Chemical stability
Avoid
Storage solution
Recommended storage temperature
Working temperature
Maximum working pressure
Flow rate
Laser label
96-Well Array Plate
Size
Material
Number of wells
200 μL
600 μL
10 mm
30 mm
5 mm
Polypropylene
All commonly used aqueous buffers, pH 1 to 14
Halogenated organic solvents, hexane
20% ethanol /1 M NaCl
2 to 8 °C
4 to 30 °C
Up to 8.0 bar g (116 psi g)
16 to 1,000 cm/hr (1)
Placed on every individual column body, and containing:
Product number, sorbent name, column volume in μL
128.3 x 86.0 x 14.0 mm
Polyoxymethylene copolymer
96
The Liquid Transfer menu of the Te-Chromu Wizard enables to set up the flow rate for each individual dispensing
and chromatography step (5.1.1.6 Liquid Transfer, Te-Chrom Wizard Software Manual, 396076, en, V1.0). Flow rate
characteristics rely on the Freedom EVO workstation characteristics, not on the RoboColumns themselves. They may
vary with another automated workstation.
(1)
www.pall.com/biopharm
3
3. Applications
ScreenExpert RoboColumns are suitable for a large variety of applications such as:
l
Parallel screening of chromatography sorbents
l
Parallel screening and optimization of chromatographic conditions in downstream process development
(conductivity, pH)
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Scale down experimental work
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Determination of dynamic binding capacity
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Determination of the optimal residence time
l
Optimization of the column regeneration procedure
4. Advice on Handling
4.1 Equipment
ScreenExpert RoboColumns are designed for robotic handling. Table 2 on page 5 is a guide for the
equipment required for robotic handling of ScreenExpert RoboColumns using the Tecan Freedom
EVO workstation.
NOTE: The workstation should hold a liquid handling (LiHa) arm equipped with stainless steel fixed tips.
4.2 Sample Preparation
We recommend loading a clarified sample on ScreenExpert RoboColumns because unclarified sample
may cause clogging of the top and bottom filter frits. Sample filtration through 0.2 μm membrane is
recommended, i.e., using Pall Acrodisc® PF Syringe Filter with hydrophilic Supor® membrane, sterile,
0.8/0.2 μm 25 mm (part number 4187).
4.3 Experimental Set Up
When using the high throughput process development approach for ScreenExpert RoboColumns,
it is suggested to use Design of Experiments (DoE) criteria for the experimental set up. This enables
to screen many different chromatographic parameters such as resin type, pH, conductivity/ionic
strength, etc., efficiently and simultaneously.
For experimental set-up and the configuration of the 96-well array plate, the Te-Chrom Wizard interface
is recommended.
4.4 Te-Chrom Wizard
The Te-Chrom Wizard is a dialog-based graphic user interface offered by Tecan and based on the
Freedom EVOwareu software.
The Te-Chrom Wizard was developed for the use of ScreenExpert RoboColumns units and provides
the possibility to configure hardware, plate layout, and the chromatographic process as well as to set
all process relevant chromatographic parameters, e.g., flow rate, volume without direct script writing.
An overview of some of the Te-Chrom Wizard functions is shown in Figure 1.
4
Table 2
Recommended Equipment for Automated Handling of ScreenExpert RoboColumns
Equipment
Details
Tips and Tricks
96-Well Array Plate
Arrange up to 96 individual
ScreenExpert RoboColumns
onto the 96-well array plate
according to application
requirements.
l
Use the Te-Chrom module (A)
to integrate the 96-well array
plate onto the robotic worktable.
The Te-Chrom module is height
adjustable and contains a waste
option (B).
l
Te-Chrom Shuttle
Arrange up to 96 individual
ScreenExpert RoboColumns
onto the 96-well array plate
according to application
requirements.
Use the robotic manipulator (RoMa) arm to transport the collection
plate from a pre-defined transfer position to any destination on the
robotic worktable, e.g., plate reader, hotels, auto sampler, etc., for
further analysis, e.g., UV measurement, ELISA, HPLC, MS, etc.
Te-Chrom Wizard
Use the Te-Chrom Wizard to
configure the chromatographic
process and to set all process
relevant parameters, e.g.,
volume, flow rate, etc.
l
Te-Chrom Module (A)
B
A
(Courtesy of Tecan)
l
l
l
l
Do not mix short (200 μL) and long (600 μL) ScreenExpert
RoboColumns units.
Make sure that ScreenExpert RoboColumns units are well clicked
into the 96-well array plate before using them.
Use the Te-Chrom module in combination with a plate stacker
Te-Stacku for the optimized handling and storage of collection
plates.
Use the waste container to drain off, e.g., conditioning,
regeneration solvents.
Make sure that the 96-well array plate is in an appropriate
distance to the collection plate before starting the experiment.
Assign labels, e.g., sorbent name and column types by using the
plate layout configurator.
Use the neutralization option to adjust the pH of the fractions after
the acidic elution of mAbs.
NOTE: Confirm suitability for every specific application with
Tecan
Plate Reader
Use an internal or external
plate reader for data evaluation
Determine the precise volume of fractions for subsequent data
plotting.
Reagent and Sample
Reservoirs
A broad spectrum of troughs,
tubes and plates can be
positioned onto the robotic
worktable by using the
appropriate Tecan carrier.
Use an 8-, 48- or 96-deep well reservoir for buffer solution /
preparation.
Collection Plate
Use any type of multi-well plate
which matches the ANSI/SDS
standard (UV- or non-UVreadable).
l
l
l
Use a half area 96-well microplate to collect fractions in a volume
range from 25 to 175 μL.
Use a standard 96-well microplate to collect fractions in a volume
range from 75 to 340 μL.
Use 96-deep well reservoirs to collect high volume fraction up to
2 mL.
www.pall.com/biopharm
5
Figure 1
Te-Chrom Wizard Functions
Hardware configuration
Plate layout
Process step editor
Liquid transfer
Plate handling
Reporting
5. Protocol
This protocol is designed as a general guideline for working with ScreenExpert RoboColumns. Adjustment
may be required depending on type of application, sample, chromatography sorbent type and ScreenExpert
RoboColumn dimension.
1. Remove cover seals
• Gently peel off the bottom seal and the row holder from the row
of eight ScreenExpert RoboColumns.
• Assemble the ScreenExpert RoboColumn units onto the 96-well
array plate. Up to 12 rows of 8 units can be assembled. Do not
mix different formats (200 and 600 μL) on the same array.
• Gently peel off the top seal of each row of 8 RoboColumn units.
2. Remove excess storage solution
• Remove remaining storage solution in the top part of the
columns by blotting the RoboColumn arrays upside down
on a soft paper tissue.
6
3. Te-Chrom – Adjusting the operating height
D
C
C
• Press and hold the fastener (C) to set the Te-Chrom to the
required height of the Te-Chrom module scale (D).
NOTE: Make sure that the ScreenExpert RoboColumn array
is as close as possible to the collection plate before starting
the experiment.
4. Te-Chrom – Loading the array plate containing ScreenExpert
RoboColumns
• Move the locking slider (E) to the left mechanical stop.
F
E
• Load the ScreenExpert RoboColumn array (F).
• Move the locking slider (E) to the right mechanical stop to
lock the RoboColumn array.
NOTE: Make sure that all RoboColumns are well fixed into
the 96-well array plate before starting the experiment.
5. Te-Chrom Wizard – Experimental set up
A. Hardware configuration
• Select the appropriate configuration of the Te-Chrom.
• Check if your Te-Chrom is equipped with a waste tray.
• Define a transfer position for the collection plate.
B. Plate layout
Click load to import an existing plate layout. To create a
new layout, follow the actions below:
• Select the ScreenExpert RoboColumn labware.
• Define the required RoboColumn matrix.
• Select the RoboColumn dimension.
• Set a desired label, e.g., resin name.
• To import or export the plate layout, click the Save button.
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C. Process configuration
• Define the chromatographic workflow by setting a desired
number of process steps.
• Use the Continuous Fraction Collection option to collect
fractions over multiple plates, without leaving gaps between
plates pipetted in different process steps and to optimize
the consumption of collection plates.
Equilibration
Load
Post Load Wash
Elution 1
Elution 2
Regeneration
Storage
D. Configure your process steps
• Edit a process step name, e.g., Equilibration.
• Check the Process Full Plate option accordingly:
– When checked: The full ScreenExpert RoboColumn array
will be processed.
– When not checked: The RoboColumn array is processed
column per column.
• Set parameters for tip washing.
• Set individual chromatographic parameters:
– Select the source of liquid.
– Set the dispensing volume.
– Set the required liquid class for aspiration.
– Set the required linear flow rate (cm/hr) or volumetric flow
rate (μL/sec).
NOTE: Calculate the required residence time (τ) by dividing
the RoboColumn volume (CV) by the volumetric flow rate (uv).
τ = CV/uv.
CAUTION! Be sure to use the correct units for uv and CV:
uv in mL/min and CV in mL, or uv in μL/min and CV in μL.
• Check the waste option accordingly:
– When checked: the waste tray will be engaged by the RoMa
(Robotic Manipulator) / LiHa (Liquid Handling).
– When not checked: the flow-through will be collected in a
desired collection plate.
• Check the Neutralization option to perform a pH adjustment
after, e.g., acidic elution of mAbs.
• Check the Data Collection option to evaluate the collection
plate in a required reader device by, e.g., UV measurement.
8
• Select the number of fractions according to the given
dispensing volume for the chromatography step if the waste
option is not checked.
• Repeat these steps to configure all chromatographic process
steps.
E. Plate handling
• Add the required amount of source plates.
• Add the required amount of destination plates.
F. Reporting
• Set output folder.
• Select output format.
After the experimental set-up, the Te-Chrom wizard will generate
a chromatographic method in Freedom EVOware software
automatically.
6. Run chromatographic method
NOTE: Make sure that all required solvents and consumables,
e.g., collection plates, are in the designated position on the
robotic worktable, before running the chromatographic method.
7. Data evaluation
• Determine the precise volume of the collected fractions by
using an appropriate plate reader.
• Determine the protein concentration by using UV measurement.
• Plot the UV data against the fraction volume to create a
pseudo-chromatogram.
NOTE: Make sure that blank values of the used solvents are
subtracted from the UV data before plotting.
14
100
90
80
70
60
50
40
30
20
10
0
12
10
ODλ = 280 nm
8. Data plotting
8
6
4
2
0
0
0.2 0.4
0.6
0.8
1.0
1.2
1.4 1.6
V (mL)
1.8
2.0 2.2
Elution Buffer (%)
NOTE: Transfer the collection plate to any location on the robotic
worktable or to secondary devices, e.g., auto sampler for further
analysis, e.g., HPLC, MS, ELISA, CGE/SDS-PAGE after path
length detection by using the RoMa.
2.4 2.6 2.8
www.pall.com/biopharm
9
9. Regeneration and storage
• For appropriate regeneration of ScreenExpert RoboColumns,
please refer to Pall recommendations for the corresponding
chromatography sorbent.
• For appropriate storage of ScreenExpert RoboColumns, please
refer to Table 1.
6. Column Scale-Up
All chromatography runs developed on ScreenExpert RoboColumns can be directly transferred onto PRC
prepacked columns (1 or 5 mL). Make sure to maintain residence time constant during scale-up.
7. Troubleshooting Guide
Fault
Possible Cause
ScreenExpert RoboColumns
are clogged
l
The sample is too viscous.
l
Increase dilution of the cell paste before lysis,
or dilute after the lysis.
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There is too much cell debris in the sample.
l
Centrifuge and/or filter the sample if unclarified
sample has been used.
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The column has been regenerated and
used too many cycles.
l
Reduce the number of cycles.
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Columns are clogged.
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See above.
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The o-ring sealing is damaged.
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Check the taught X- and Y coordinates for
ScreenExpert RoboColumn labware for
precision.
l
The insertion depth of the robotic needle
is incorrect.
l
Check the insertion depth (16 mm from the
top of the column) of the robotic needle.
ScreenExpert RoboColumns
are lifted up from the 96-well
array plate during the pull out
of the robotic needles
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ScreenExpert RoboColumns are not well
fixed into the 96-well array plate.
l
Make sure that all ScreenExpert RoboColumns
are well fixed into the 96-well array plate before
starting the experiment.
Low protein recovery in the
elution fraction(s)
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The residence time for sample loading is
too low.
l
Decrease the flow rate for sample loading.
l
The choice of the sorbent and/or process
conditions is not appropriate.
l
Check and adapt if needed the sorbent and/or
process conditions.
ScreenExpert RoboColumns
are leaking
10
Action
8. Ordering Information
Description
Part Number
ScreenExpert RoboColumn MEP HyperCel 200 μL, row of 8
ScreenExpert RoboColumn MEP HyperCel 600 μL, row of 8
ScreenExpert RoboColumn HEA HyperCel 200 μL, row of 8
ScreenExpert RoboColumn HEA HyperCel 600 μL, row of 8
ScreenExpert RoboColumn PPA HyperCel 200 μL, row of 8
ScreenExpert RoboColumn PPA HyperCel 600 μL, row of 8
ScreenExpert RoboColumn HyperCel STAR AX 200 μL, row of 8
ScreenExpert RoboColumn HyperCel STAR AX 600 μL, row of 8
ScreenExpert RoboColumn CM Ceramic HyperD® F 200 μL, row of 8
ScreenExpert RoboColumn CM Ceramic HyperD F 600 μL, row of 8
ScreenExpert RoboColumn HyperCel STAR CEX 200 μL, row of 8
ScreenExpert RoboColumn HyperCel STAR CEX 600 μL, row of 8
ScreenExpert RoboColumn CMM HyperCel 200 μL, row of 8
ScreenExpert RoboColumn CMM HyperCel 600 μL, row of 8
ScreenExpert RoboColumn KANEKA KanCapAu 200 μL, row of 8
ScreenExpert RoboColumn KANEKA KanCapA 600 μL, row of 8
96-well RoboColumn array plate
SR2MEP
SR6MEP
SR2HEA
SR6HEA
SR2PPA
SR6PPA
SR2STARAX
SR6STARAX
SR2CMCHDF
SR6CMCHDF
SR2STARCEX
SR6STARCEX
SR2CMM
SR6CMM
SR2KANCAPA
SR6KANCAPA
SR96WAP
Complementary Products
PRC Prepacked Columns
PRC Column 5x50 MEP HyperCel, 1 mL
PRC Column 8x100 MEP HyperCel, 5 mL
PRC Column 5x50 HEA HyperCel, 1 mL
PRC Column 8x100 HEA HyperCel, 5 mL
PRC Column 5x50 PPA HyperCel, 1 mL
PRC Column 8x100 PPA HyperCel, 5 mL
PRC Column 5x50 HyperCel STAR AX, 1 mL
PRC Column 8x100 HyperCel STAR AX, 5 mL
PRC Column 5x50 Q Ceramic HyperD F, 1 mL
PRC Column 5x50 CM Ceramic HyperD F, 1 mL
PRC Column 5x50 HyperCel STAR CEX, 1 mL
PRC Column 8x100 HyperCel STAR CEX, 5 mL
PRC Column 5x50 CMM HyperCel, 1 mL
PRC Column 8x100 CMM HyperCel, 5 mL
PRC Column 5x50 KANEKA KanCapA, 1 mL
PRC Column 8x100 KANEKA KanCapA, 5 mL
PRC05X050MEPHCEL01
PRC08X100MEPHCEL01
PRC05X050HEAHCEL01
PRC08X100HEAHCEL01
PRC05X050PPAHCEL01
PRC08X100PPAHCEL01
PRCSTARAX1ML
PRCSTARAX5ML
PRC05X050QCHDF01
PRC05X050CMCHDF01
PRCSTARCEX1ML
PRCSTARCEX5ML
PRCCMMHCEL1ML
PRCCMMHCEL5ML
PRC05X050KANCAPA
PRC08X100KANCAPA
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11
8. Ordering Information (continued)
Description
Part Number
AcroPrep™ ScreenExpert Plates
MEP, HEA and PPA HyperCel Mixed-Mode Sorbents
MEP HyperCel Mixed-Mode Sorbent
HyperCel STAR AX Anion Exchange Sorbent
96WP3MIXM50
96WPMEP50
96WPSTARAX50
Multi-Well Plates
AcroPrep Advance 96-Well Filter Plate
8129
AcroPrep Advance Filter Plate for Ultrafiltration
350 μL, Omega™ 3K MWCO (10/pkg)
350 μL, Omega 10K MWCO (10/pkg)
350 μL, Omega 30K MWCO (10/pkg)
350 μL, Omega 100K MWCO (10/pkg)
8033
8034
8035
8036
Syringe Filter
Acrodisc PF Syringe Filter with hydrophilic Supor membrane, sterile, 0.8/0.2 μm 25 mm
4187
Visit us on the Web at www.pall.com/biopharm
E-mail us at biopharm@pall.com
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Port Washington, NY, USA
+1 800 717 7255 toll free (USA)
+1 516 484 5400 phone
biopharm@pall.com e-mail
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+41 (0)26 350 53 00 phone
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International Offices
Pall Corporation has offices and plants throughout the world in locations such as:
Argentina, Australia, Austria, Belgium, Brazil, Canada, China, France, Germany,
India, Indonesia, Ireland, Italy, Japan, Korea, Malaysia, Mexico, the Netherlands,
New Zealand, Norway, Poland, Puerto Rico, Russia, Singapore, South Africa, Spain,
Sweden, Switzerland, Taiwan, Thailand, the United Kingdom, the United States, and
Venezuela. Distributors in all major industrial areas of the world. To locate the Pall
office or distributor nearest you, visit www.pall.com/contact.
The information provided in this literature was reviewed for accuracy at the time of
publication. Product data may be subject to change without notice. For current
information consult your local Pall distributor or contact Pall directly.
© 2015, Pall Corporation. Pall,
, Acrodisc, AcroPrep, HyperCel, HyperD, Omega
and Supor are trademarks of Pall Corporation. Filtration.Separation.Solution. is a
service mark of Pall Corporation. ® indicates a trademark registered in the USA
and TM indicates a common law trademark. u MediaScout and RoboColumn are
trademarks of Atoll GmbH. Freedom EVO, Freedom EVOware, Te-Chrom and
Te-Stack are trademarks of Tecan Group. KANEKA KanCapA is a trademark of
KANEKA Corporation.
1/16, GN16.9896
USD 2937(c)