Easy Set-Up for Protein Crystallization Experiments
App Note / Case Study
Published: October 31, 2023
X-Ray analysis of protein crystal has contributed to major advances in the determination of protein structures.
However, successfully growing protein crystals is challenging and can be impacted by factors such as the nature of the reagents, the pH and the temperature.
This app note highlights how a handheld electronic pipette can help researchers save time and increase homogeneous crystal growth.
Download this app note to learn how to achieve:
Fast and easy transfer of liquids to crystallization plates for high -throughput screening of 96 crystallization conditions
Increased pipetting precision when pipetting into reservoir or crystallization wells
More robust crystal growth due to fast set-up of crystallization reactions
Easy set-up for high throughput protein
crystallization experiments using the
VIAFLO 96/384 handheld electronic pipette
Protein crystallization and subsequent X-ray analysis have
contributed to major advances in the determination of
protein structures. Successfully growing protein crystals is
challenging and depends on many factors, such as the nature
of the reagents, the pH and the temperature. The sitting drop
vapor diffusion technique is a protein crystallography method
that is very well suited to a 96 well crystallization plate format.
• Accommodate and process 96 well crystallization plates
on the VIAFLO 96/384 for high throughput crystallization
• Increased pipetting precision when pipetting into reservoir
or crystallization wells.
• The VIAFLO 96/384 handheld electronic pipette allows
easy and rapid set-up of up to 96 different crystallization
conditions for increased productivity and reproducibility.
The following protocol describes a set-up for performing
an initial protein crystallization screen using the VIAFLO
96/384 handheld electronic pipette together with a three
position stage (Figure 1).
In this protocol, a 96 deep well block filled with 96
different reagents (Morpheus® HT-96, Molecular
Dimensions), placed on Position A, is used as the source
for a 96 condition crystallography screen. The protein
crystallography plate (3 Lens Crystallisation Microplate
(UVXPO), SWISSCI) is placed on Position AB, while
tips and the tip waste are accommodated on Position
B. Three different protein solutions are filled into PCR
strips and accommodated on a PCR cooling block, which
is exchanged with the Morpheus HT-96 screen plate as
Three position stage
All wells of a plate of this type can be filled simultaneously
with perfect tip alignment using the VIAFLO 96/384 handheld
electronic pipette with a 96 channel pipetting head. Using
a single aspiration step reduces the time to cover the entire
well plate, which helps to prevent evaporation of precious
samples, resulting in more homogeneous crystal growth
across the plate.
• Optimal pipetting heights for transfer of proteins and
reagents directly into the crystallization plate are
• The VIAFLO 96/384 allows for fast set-up of crystallization
reactions, which helps to avoid evaporation effects,
resulting in more robust crystal growth.
1. Transfer of the screening
STEP: Transfer the
agents into each well of the
HOW TO: Load tips and launch the program ‘CRYST_
SCREEN’ to fill the lens and reservoir wells (Figure 2) of the
crystallization plate with the crystallization screening agents.
Aspirate 40 µl of the Morpheus HT-96 reagent and dispense 2
µl back into the 96 deep well block (Position A), as indicated
in the pipetting program. Dispense 2 µl into each lens well
(1-3), and 30 µl into the reservoir well (4), of the crystallization
plate on Position AB according to Figure 2. Move the plate
slider of the three position stage to access the different wells
of the crystallization plate more easily. Discard the pipette
tips with the remaining liquid. The accuracy of these pipetting
steps is increased by using a pre- and post-dispense.
• Pipetting heights are defined in the program to prevent the
tips from touching the bottom of the plate.
• The volumes dispensed can be easily adjusted to allow
investigation of different ratios of protein and precipitant.
Figure 1: The VIAFLO 96/384 handheld electronic pipette with a three position stage showing the experimental set-up. Position A: crystallization screen plate.
Position AB: crystallization plate. Position B: 125 µl Sterile, Filter, Low Retention GripTips and waste to collect the discarded tips.
Deck Position A
Crystallization screen plate and
INTEGRA cooling block with PCR strips
filled with the different proteins
Deck Position AB
Deck Position B
125 µl Sterile, Filter, Low Retention
GripTips and waste
A 96 channel pipetting head (2-50 µl) is used together with 125 µl Sterile, Filter, Low Retention GripTips. Customized VIALINK
programs are provided to set up a protein crystallization screen with the VIAFLO 96/384 handheld electronic pipette.
Figure 2: Pipetting scheme of the crystallization plate well. Lens wells: 1-3. Reservoir well: 4.
Figure 3: All four wells (three lens wells and one reservoir well) of the crystallization plate are accessible with the VIAFLO 96/384 handheld electronic pipette.
2. Transfer of protein
STEP: Transfer up to three
different protein solutions into
the wells of the crystallization
HOW TO: 30 µl of each of the three different protein solutions
are manually distributed into 8 tubes of a PCR strip to allow
subsequent aspiration with 8 pipette channels in parallel. The
PCR strip containing protein solution 1 is positioned in column
12 of a cooling block, which acts as a support. Instead of loading
all 96 tips at once, load 8 tips to the last column of the pipetting
head and aspirate 28 µl of protein solution 1. Perform a predispense (2 µl) and then transfer 2 µl to lens well 1 (Figure 2)
of each column of the crystallization plate (Figure 4). Perform
a post-dispense (2 µl). Discard the tips and exchange the PCR
strip for one containing protein solution 2. Transfer the protein
solution into lens well 2. Repeat the process for protein solution
3 and lens well 3.
• Aspiration and dispense speeds can be easily adapted to
allow optimized handling of the protein solutions.
• The column detect mode of the VIAFLO 96/384 handheld
electronic pipette offers users haptic feedback for accurate
and easier guidance when moving to the next column.
• Protein solutions can be kept chilled in the cooling block if
• Protein solutions can also be stored in a 12 column reagent
reservoir if higher volumes of the protein solution are
available. The corresponding VIALINK program can be
found in the download section.
• Alternatively, 2 µl of protein solution can be transferred from
a PCR strip into the crystallization wells using a VOYAGER
8 channel 12.5 µl adjustable tip spacing electronic pipette
with 12.5 µl Sterile, Filter, Low Retention GripTips.
Figure 4: Pipetting and transfer of the protein solution with the VIAFLO 96/384 handheld electronic pipette.
3. Sealing of the
STEP: Seal and incubate the
• The duration and temperature of sitting drop vapor diffusion
depends on your proteins.
• The VIAFLO 96/384 handheld electronic pipette
allows transfer of liquids to crystallization plates in a
fast and easy way for high throughput screening of 96
• Optimal consistency and reproducibility of the results are
achieved by using the VIAFLO 96/384 to simultaneously
transfer 96 liquids.
• The protocol for different protein solutions and
crystallization screens can be easily adapted using the
• Crystallization plates for sitting drop vapor diffusion in the
ANSI/SLAS plate format are optimal for processing with
the VIAFLO 96/384 handheld electronic pipette.
Manufacturer Part Number Description Link
INTEGRA Biosciences 6001/6031 VIAFLO 96 or 384 handheld
electronic pipette (base unit)
INTEGRA Biosciences 6230 Three position stage for 96 and
384 well plates
INTEGRA Biosciences 6106 96 channel pipetting head 50 µl https://www.integra-biosciences.com/global/en/
INTEGRA Biosciences 6362 12 Column Reagent Reservoir
INTEGRA Biosciences 6565 125 µl Sterile, Filter, Low
INTEGRA Biosciences 6250 PCR 96 Well Cooling Block https://www.integra-biosciences.com/global/en/
CORNING CLS 3741 Corning® Thermowell GOLD PCR
8 well tube strips
SWISSCI UVXPO-3LENSLP 3 Lens Crystallisation Plate https://swissci.com/wp-content/uploads/2020/03/3-
DIMENSIONS MD1-47 Morpheus HT-96 https://www.moleculardimensions.com/products/
INTEGRA Biosciences AG
7205 Zizers, Switzerland
T +41 81 286 95 30
F +41 81 286 95 33
INTEGRA Biosciences Corp.
Hudson, NH 03051, USA
T +1 603 578 5800
F +1 603 577 5529
INTEGRA Biosciences Ltd.
Egham, Surrey TW20 9EY, UK
INTEGRA Biosciences SAS
95062 Cergy-Pontoise Cedex 1, France
T +33 (0)1 34 30 76 76
F +33 (0)1 34 30 76 79
INTEGRA Biosciences Deutschland GmbH
35444 Biebertal, Deutschland
T +49 6409 81 999 15
F +49 6409 81 999 68
Brought to you by
Download the Application Note for FREE Now!
Information you provide will be shared with the sponsors for this content. Technology Networks or its sponsors may contact you to offer you content or products based on your interest in this topic. You may opt-out at any time.