2011 course on Molecular Diagnostic Automation - Part 1 - Nucleic Acid Extraction. This is from early 2011. Prices and Specifications of instruments may have changed. Part 1 of 3

1. Patrick Merel
Biomedical Innovation Platform (PTIB), Pessac, France
pmerel@mac.com
Nucleic Acid Extraction Automation
in Molecular Diagnostic
part 1
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2. Nucleic Acid (NA) extraction in molecular diagnostic
DNA and RNA from...
Blood, Plasma, Cells … Body Fluids
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3. NA Extraction Steps Automation
Quality of DNA/RNA is crucial for the PCR, RT/PCR steps
Extraction procedures often require centrifugation or filtration
Which are difficult steps to automate
Or, which require complex instrumentation
However, there are now numerous options for automation of NA extraction steps
Semi-automated protocols (microtiterplate based)
Automated protocols on robotic workstations
Dedicated and automatic workstations
But first, let’s go shopping for the perfect NA extraction technology...
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4. NA organic extraction
Organic extraction is a conventional technique that uses
organic solvents to extract contaminants from cell lysates.
The cells are lysed using a detergent, and then mixed with
phenol, chloroform, and isoamyl alcohol.
The correct salt concentration and pH must be used during
extraction to ensure that contaminants are separated into the
organic phase and that DNA remains in the aqueous phase.
DNA is usually recovered from the aqueous phase by alcohol
precipitation.
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5. NA Phenol-chloroform extraction
Equal volumes of a phenol:chloroform mixture and the aqueous DNA
sample are mixed, forming a biphasic mixture.
The proteins will partition into the organic phase while the DNA (as well as
other contaminants such as salts, sugars, etc.) remain in the aqueous
phase.
It is then followed by precipitation with ethanol, as ethanol precipitates the
nucleic acid.
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6. Nucleic Acid Extraction by centrifugation
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7. DNA extraction by Salting-Out
•Salting-out is another conventional technique where proteins
and other contaminants are precipitated from the cell lysate using
high concentrations of salt such as potassium acetate or
ammonium acetate.
•The precipitates are removed by centrifugation, and the DNA is
recovered by alcohol precipitation.
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8. DNA Separation by Silica Adsorption
•This method is based on the selective adsorption of nucleic acids to a silica-gel
membrane in the presence of high concentrations of chaotropic salts.
•Use of optimized buffers in the lysis procedure ensures that only DNA is
adsorbed while cellular proteins, and metabolites remain in solution and are
subsequently washed away.
•Ready-to-use DNA is then eluted from the silica-gel membrane using a low-salt
buffer.
•No alcohol precipitation is required but filtration or combination of filtration
+centrifugation procedures are needed.
Most nucleic acid purification techniques that use silica in combination with chaotropic reagents such as guanidine hydrochloride are covered by U.S.
Patent No. 5,234,809, European Patent No. 0,389,063, and equivalents in other countries. It is commonly known as the "Boom" patents and is owned
by Biomerieux.
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9. Silica beads based NA extraction
For DNA
or RNA
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10. Centrifugation + filtration NA extraction
Mainly single tube based,
but various options for
microtiterplate processing are now
available
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11. Filtration + centrifugation NA extraction
Filtration only procedures available
for single unit processing, and
microtiter plate based products for
robotic handling.
Most of the time filtration
procedures end with a centrifugation
step.
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12. Magnetic beads based NA extraction
High quality nucleic acids
Scalable
No centrifugation
No vacuum steps
Rapid purification
Reproducible
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13. Dynal Dynabeads Technology
•D y n a b e a d s a r e s u p e r p a r a m a g n e t i c ,
monosized polymer particles, providing a
consistent, defined surface for the adsorbtion or
coupling of various bioreactive molecules.
•The beads are magnetisable and super
paramagnetic, meaning they are only magnetic
in a magnetic field.
•Due to this property, the beads can easily be
resuspended when the magnetic field is
removed.
•The technology behind the beads was licensed
to Dyno Industrier in 1980, which later formed
Dynal Biotech together with Apothekernes
Laboratorium in 1986.
Scanning Electron Microscopic pictures of the
monosized Dynabeads® (A) and alternative
magnetisable particles from other suppliers (B-E).
Courtesy of Mikal Heldal, University of Bergen,
Norway. 13
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14. Dynal Dynabeads Procedure
•Chaotropic soln (GTC) + silica magnetic particles
•Chaotropic solution, removes non-bound matter
•EtOH, removes GTC
•Water, removes EtOH
•Water or low salt buffer
๏Lysis and binding 5+5 min
๏Wash, Rinse 3x1 min
๏Elution 30 s RT
๏or 5 min 65C
๏Total @ RT <15 min
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15. Agencourt SPRI Technology
•The Agencourt SPRI (Solid Phase Reversible
Immobilization) technology is a patented purification
technology developed at MIT's Whitehead Institute
(Hawkins, et. al., Nucleic Acids Res. 1995 (23):
4742-4743).
•Nucleic acids are immobilized onto paramagnetic
microparticles (1µM magnetic carboxylate-modified base
particles) using specific buffer conditions. Flexibility is
added to the system through modification of the binding
buffer, which alters the type and size of immobilized
nucleic acid.
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16. Agencourt SPRI Technology
Lysis 30 min RT
Bind 5 min
Capture 15 min
Washing1 2x10 min
Washing2 2x6 min
Capture+Elution 2+10 min
Total @ RT 100 min
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17. Magnetic Extraction Pros and Cons
Pros
•Usually very fast procedures
•Robotic friendly
•Rapid growing segment for instrumentation
•Cheaper ?
•Potential for specific extraction
Cons
•Manipulation of small volume samples
•For small to medium quantity of DNA
•Magnetic stand/plate needed
•Usually generic extraction
•Not all procedures equal in regards of time requirement
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18. NA extraction by hybridization
Different companies have developed over the past years, specific hybridization capture
procedures:
RNAture (Hitachi Chemical), Xtrana, ProLinx, Gen-Probe, DNA Research Innovation
(DRI now part of Invitrogen/Life Technologies) have been the most visible.
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19. Gen-Probe Target Capture Technology
Target Capture
•Maximizes assay sensitivity & specificity. •Allows for simplified specimen processing.
•RNA target is “captured” with magnetic •Patented process with specific probes
beads.
coated magnetic particles.
•Eliminates inhibitors in swab and urine
samples.
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20. Invitrogen ChargeSwitch Technology
ChargeSwitch gDNA
50-100 µl
Lysis 1 min RT
Capture 1 min
Washing 3x1 min
Elution 1+4 min
Total @ RT 10 min
How it works
ChargeSwitch Technology provides a switchable surface charge that is dependent on the pH of
surrounding buffer to purify nucleic acids (Figure 1). In conditions of pH < 6.5, the CST surface has a
positive charge that selectively binds the negatively charged nucleic acid backbone. Proteins and other
contaminants are removed in an aqueous wash buffer. For elution, the charge on the CST surface is
switched off by raising the pH to 8.5. Purified nucleic acid elutes instantly and is ready for
downstream use.
Advantages of the ChargeSwitch Technology
• Novel, pH dependent switch action
• Avoids introduction of ethanol, organic solvents or chaotropic salts
• Improved downstream performance for high specification applications
• Unmatched high sensitivity for micro samples 20
20

21. Very recent new approach for NA extraction
Enzyme based NA extraction automated on the epMotion 5075MC
DNA extraction using a thermostable proteinase.
…hands off, closed tube and all you need is a thermal cycler.
WorkFlow
Zygem: PrepGEMand ForensicGEM
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22. Very recent new approach for NA extraction
Boreal Genomics instruments are based on SCODA: a •Up to 100X more efficient for contaminant rejection
novel, proprietary electrophoresis technology that •High yield even in very dilute samples
selectively concentrates nucleic acids from large sample •Simple operation
volumes to a small, concentrated amount of gel or buffer. •Target length selection during extraction
Advantages:
•Protection of high molecular weight targets
•Potential for sequence specific extraction including
short RNA/DNA fragments
This is accomplished by
means of rotating electric
fields that act uniquely on
long, charged polymers
and leave other molecules
unaffected.
1x5mL or 4x1 mL input
• Target run times < 30 min
• Digital high voltage
control
• Unattended operation
• Lysate to purified sample
in one step
• Extraction in 10 – 50 uL of
buffer
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23. NA extraction Automation: why?
Time consuming step in MDx
Sample throughput
Hand on time
Reproducibility
Standardization
Cost
Instrumentation availability
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24. NA extraction automation options
✓Microtiter Plate based Procedures
✓Generic Robotic Instrumentation
✓Dedicated Robotic Instrumentation
✓In Vitro Diagnostic Robotic Instrumentation
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25. Microtiterplate based NA extraction
•Qiagen and Macherey-Nagel have been pioneers in providing DNA
extraction protocols on 96 well-plates
Qiagen, http://www.qiagen.com
DNAeasy, RNAeasy, QIAamp, QIAfilter
Macherey-Nagel, http://www.macherey-nagel.com
NucleoBond, NucleoSpin, NucleoTrap, Multi-96
•Additional companies providing 96 well-plate compatible kits for DNA/RNA
extraction:
Promega, BioLine, Stratagene, Millipore, Ambion, Invitrogen…
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26. High Throughput DNA extraction with Macherey-Nagel
The NucleoSpin Multi96 blood kit
from macherey-Nagel allows rapid
parallel DNA extraction from blood,
serum, plasma, cell cultures, buffy
coats.
Features:
silica membrane technology
complete removal of PCR inhibitors
no organic solvents
200 µl whole blood or 107 lympho. or
cultured cells can be processed
Simultaneous processing of 192
samples
2 x 96 well plates
or
n x 8 strips on a 96 deep-well block
Processing time >2hrs
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27. Microtiterplate compatible viral DNA extraction
Bench top centrifuge for
Nucleospin multi 96
DNA extraction.
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28. HighThroughput DNA extraction from
centrifugation to filtration
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29. High Throughput DNA extraction from centrifugation
to filtration
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30. Semi-Automation of the QIAamp 96 Spin Blood Kit on the
Biomek 2000
Setup of the Beckman
Coulter Biomek 2000
Tablet
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31. Qiagen QIAamp 96 Spin Blood Kit setup on the Biomek 2000
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32. Semi-Automation of the QIAamp 96 Spin Blood Kit on the Biomek
2000
Centrifugation of the Centrifugation of the
Filtration Unit + Filtration Unit +
Disposable Deep-Well Storage Deep-Well
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33. NA extraction automation options
✓Microtiter Plate based Procedures
✓Generic Robotic Instrumentation
✓Dedicated Robotic Instrumentation
✓In Vitro Diagnostic Robotic Instrumentation
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34. Generic Robotic Instrumentation
✓Develop your own procedure
based on manual protocols
some are robotic friendly
✓Follow reagents providers recommendations
✓Follow robotic instrument providers recommendations
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35. Generic Robotic Workstations
with available molecular testing applications.
Company Instrument IVD-D or CE-IVD NA extraction PCR applications Web Address
compliant; comments applications available
available
Agilent Automation Biocell, Bravo, No; very high
Yes Yes chem.agilent.com/
Solutions BenchCel throughput
Versa serie, Versa
Aurora Instruments no Yes Yes aurora-instr.com/
mini
Biomek 2000,3000
Beckman Coulter, Inc. No Yes Yes beckmancoulter.com/
Biomek NX,FX
SciClone ALH3000
No Yes Yes
Caliper LifeSciences caliperls.com/
Zephyr No Yes Yes
Twister II+AB7900 No No Yes
CyBi-Dilus, -
Cybio No Yes Yes cybio-ag.com/
RoboSpense
epMotion 5070
No; MC with
Eppendorf epMotion 5075 Yes Yes eppendorf.com/
integrated PCR
LH, VAC, MC, TMX
STAR, STARlet, Yes; NAT pooling
Hamilton Yes Yes hamiltoncomp.com/
Nimbus system
No; very high
Labcorp-Protedyne BioCube, Radius Yes Yes protedyne.com/
throughput
Janus CS Autoplex,
Perkin-Elmer No Yes Yes las.perkinelmer.com/
Forensic
Tecan Freedom Evo Yes Yes Yes tecan.com/
No; very high
ThermoCRS CRS DNA Yes Yes thermo.com/
throughput
Xiril Xiril 75, 100, 150 No Yes Yes xiril.com/
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36. ThermoCRS Extraction platforms
High throughput (up to 14, 96-well microplates/hour)
Proven reliability
Uses well-established DNA purification protocols
Adaptable to any magnetic bead based
separation method
Unattended 24 hour operation and remote monitoring
POLARA™ Open Architecture Laboratory Automation
Software
The CRS DNA Purification System is an automation tool for bead
based methods using magnetic particles. This includes binding of DNA
to magnetic beads, washing of the bound material, elution from the solid
phase and transfer to a final destination plate.
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37. Agilent Automation Solutions-Bravo and BioCel
BioCel
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38. Agilent Automation Solutions: Bravo and BenchCel
A flexible workstation providing high-throughput gDNA
isolation
• Up to 10 microplates can be processed in one run without
user intervention using the Invitrogen Charge Switch
EasyPlex procedure
• Microplate processing time is approximately 10 min per
microplate (excluding incubation time ~ 30min)
Bravo
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39. Protedyne BioCube Platforms
Protedyne BioCubes
LX, SX, EX
Performance: Approximately
7,000 DNA samples can be
prepared per eight-hour shift
using a Protedyne 96-well,
vacuum-based plasmid DNA
purification procedure.
Also, QiaAmp Genomic DNA on
the BioCube: 14 plates per 8
hour shift
Protedyne Radius
Protedynes bench-top robotic a protocol for the Invitrogen ChargeSwitch
system EasyPlex gDNA kit using the gDNA isolation
workstation in which up to 10 microplates (96-
well) can be processed simultaneously.
In May 2007, Protedyne became a wholly owned subsidiary of Laboratory Corporation of America. 39
39

40. Hamilton Robotics Platforms
STARlet STAR STARplus
Nimbus MICROLAB 4000 MICROLAB 4200
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41. Tecan Extraction Platforms
T h e M i n i P re p , f ro m D r u g The Genesys serie with four or eight tip arm for
Discovery (mother - daughter plate The Freedom Evo family from
high throughput. NA extraction automation has
replication), to Molecular Biology 1m to 2m deck size. A special
been fully automated using the magnetic separator
(Sequencing preparation, PCR mention for the IVD-D compliant
MagS and solid phase extraction through columns.
amplification assays) and Freedom Evo Clinical.
The extraction is usually performed in 96 well
Diagnostics (Immunophenotyping, cartridge blocks. These are placed on vacuum
ELISA Assays). chambers on the instrument worktable and can be
evacuated by software controlled vacuum pump
VacS.
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42. Invitrogen reagents automation and large volumes procedures
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43. Xiril Robotic Platforms
X75 X100
Xiril Robotic Workstations have a 1-tip, 2-tip or 4-tip liquid handling arm with selectable
and exchangeable pump volumes. Each pump is equipped with a tip adaptor that fits
disposable tips with or without filters.
For added flexibility and increased sample throughput, the Xiril X100 and X150 can be
configured with an additional liquid handling arm for specific Life Science applications.
Macherey, Agencourt, Promega application notes available.
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44. Beckman Coulter Biomek Platforms
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45. Beckman Coulter NX and Biomek Platforms
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46. Robotic Workstation Industrials “Partners”
Beckman Coulter Hamilton Thermo-Jouan- CRS PerkinElmer Tecan Self-Contained units
Robotics
Biomek 2000, Biomek 3000 MicroLab, Starplus, CRS DNA Purification MultiProbe IIEX, Genesis RSP 100, 150, Agilent Biocel, Bravo,
Starlet System DNAtrack, Janus 200 Protedyne BioCube
Evo
Biomek FX, Biomek NX AutoSprint MiniPrep 60 MolBio
Multimek
Promega, Xtrana, Millipore, Agowa, Roche BilaTec Agencourt, Agowa, Qiagen, Macherey-Nagel, Various, Invitrogen
Agencourt, Qiagen Macherey-Nagel, Agowa
Promega
Filtration, Magnetic, Filtration, purification Magnetic Filtration, Magnetic Filtration, Magnetic all types
hybridation
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47. Robotic friendly NA extraction reagents
๏ Automated Protocols ๏ Automated Wizard® Magnetic 96 DNA Plant
System: Scaled Up Protocol for Increased DNA
Yield
๏ Automated SV 96 Total RNA Isolation System
Protocol
๏ Automated Wizard® MagneSil® Sequencing
Reaction Clean-Up System Protocol for 96-
Well Plates
๏ Automated Wizard® Magnetic 96 DNA Plant
System Protocol
๏ Automated MagneSil® ONE, Fixed Yield Plant
Genomic DNA Purification Protocol
๏ Automated Wizard® SV 96 Genomic DNA
Purification System Protocol
๏ MagneSil® KF, Genomic System
๏ Automated Wizard® SV 96 PCR Clean-Up
System Protocol
๏ Automated Wizard® MagneSil™ 384-Well PCR
Clean-Up System Protocol
๏ Automated Wizard® MagneSil™ 384-Well
Sequencing Reaction Clean-Up System Protocol
๏ Automated Wizard® MagneSil® 96-Well PCR Example of Promega’s reagents
Clean-Up System Protocol
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48. NA extraction automation options
✓Microtiter Plate based Procedures
✓Generic Robotic Instrumentation
✓Dedicated Robotic Instrumentation
✓In Vitro Diagnostic Robotic Instrumentation
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48

49. Dedicated Workstations For NA extraction 1/4
Applied BioSystems ABI Prism 6100 Filtration 96 96/30min jusqu’à 1m de ARN, ADN appliedbiosystems.com
cellules
ABI Prism 6700 Filtration; archivage, 96 96/75min jusqu’à 1m de ARN, ADN
préparation PCR cellules
Autogen AutoGenflex 3000 Centrifugation 48 48/4-6hr jusqu’à 5mL ADN, ARN, autogen.com
plasmides,
cosmides
AutoGenPrep 2000 48 48/4-6hr 1mL ADN, tissus,
plasmides,
cosmides, levures,
plantes
AutoGenPrep 245 24 24/3.5hr 700µL ARN, ADN, tissus,
plasmides
AutoGenPrep 965 4x96 384/4-6hr Jusqu’à 250µL ou ADN, plasmides,
1mL cosmides
Beckman Coulter, Vidiera NsP Filtration; 96 96/2.5hr Jusqu’à 300µL ADN, ADN et ARN beckman.com
Inc. quantification, viral
normalisation,
préparation PCR
SPRI-TE Magnétique/ 10 10/30min NS ADN
Magtration; cartouches
de réactifs
Bee Robotics QuatroProbe Magnétique 48 NS 200µL à 1mL NS beerobotics.com
Bioer Gene-Pure NPA-32 Magnétique/Magnetic 32 NS 50 à 800µl ADN, ARN bioer.com.cn
rod
Biomérieux Extractor Méthode de Boom 10 120/8hr ADN 100µL, jusqu’à ADN, ARN biomerieux.com
2mL
EasyMag Magnétique/Méthode 24 60min Jusqu’à 1mL
de Boom
Bioneer HT-Prep Pro Filtration 96 60min Jusqu’à 300µL ADN, ARN bioneer.co.kr
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50. Dedicated Workstations For NA extraction 2/4
Chemagen Chemagic Magnétique/Magnetic 12, 96 150/8h 10mL 10µL à 10mL DNA, RNA, RNAv, chemagen.de
rod (45min/12), mRNA
4000/8h 1.5mL
(15min/96)
Prepito 1-12 30min 10µL à 600µL
Corbett LS part of QIAxtractor FIltration 8-96 8/45min, 96/96min 180 à 200µL DNA, RNA, RNAv, corbettlifescience.com
Qiagen mRNA
Eppendorf epMotion 5075 VA, Filtration 1-96 NS NS DNA, RNA, tissus, eppendorf.com
TMX plasmides
FujiFilm QuickGene-610 Filtration sur 6 12min 2mL ADN fujifilm.com
QuickGene-810 Membrane poreuse de 8 6min 200µl ADN, ARN, tissus,
80µM plasmides
QuickGene-mini80 8 6min (manuel) 200µl ADN, ARN, tissus,
plasmides, cellules
GeneSystems GeneExtract Filtration 6 Semi-automatique > 1L de liquide ADN genesystems.fr
Genomic ExtraGene Centrifugation 48 240/jour 0.5 à 10mL ADN, ARN, Proteines genomics-tools.com
HandyLab part of BD Lynx Magnétique; 24 35min NS ADN, ARN handylab.com
cartouches de réactifs
Invitrogen iPrep Magnétique/ 12 26min 350µL ADN invitrogen.com
Magtration; cartouches
de réactifs
Magnetic Magnatrix 1200 Magnétique/Magnatrix 12-96 20min/12 NS ADN, ARN magbio.com
Biosolutions AB
Magnatrix 8000plus 8-192 NS NS
Mole GeneMole Magnétique; 16 NS 50µL à 1mL ADN, ARN, cellules, molegenetics.com
cartouches de réactifs tissus
Nordiag Arrow Magnetic, Bug’s and 1-12 30min 200-500µL DNA, RNA nordiag.com
Beads
Bullet 1-96 1.5hr 20 à 200µL DNA, RNA
Promega Maxwell 16 Magnétique/Magnetic 16 30min 50 à 400µL ADN www.promega.com
rod; cartouches de
réactifs
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51. Dedicated Workstations For NA extraction 3/4
Precision System Mag. Syst. 6Mx Magnétique/ 6-48 NS 100 à 1000µl ADN, ARN pssbio.com
Science Magtration; cartouches
de réactifs
Mag. Syst. 8Lx Magnétique/ 1-8 50min 1-10mL ADN
Magtration; cartouches
de réactifs
Magt. Syst.12GC Magnétique/ 1-12 30min 100 à 200µL ADN, ARN, ADN et
Magtration; cartouches ARN viral, tissus,
de réactifs plasmides
SX-6G Magnétique/Magtration 48 80min 25 à 500µL ADN, ARN
SX-8G Magnétique/Magtration 96 80min 5 à 40µL ADN, ARN
SX-96GC Magnétique/Magtration 4x96 NS 10 à 40µL ADN, ARN
Mag. Syst.Ba-X Magnétique/Magtration 1-3 0.2ml,1ml, 3ml ADN, ARN
Qiagen BioRobot Universal System
Filtration + 4x96 4x96/2-3hr jusqu’à 200µL ADN, ADN et ARN qiagen.com
centrifugation + viral, plasmides
Magnétique; PCR
setup
BioRobot MDX DSP** Filtration + 4x96 4x96/2-3hr jusqu’à 200µL ADN, ADN et ARN
centrifugation + viral
Magnétique ; PCR
setup; CE IVD-D
EZ1, EZ1 advanced, Magnétique/ 6, 6, 14 20min 200 à 350µL ADN, ADN et ARN
EZ1 advanced XL Magtration; cartouches viral, tissus
de réactifs; CE-IVD-D
BioRobot 48/96 Magnétique/Magtration 0,5 48/2hr 100 à 300µL ADN, ADN/ARN,
ARN, mARN
BioSprint 15/96 Magnétique/Magnetic 0,15625 20min 10µL à 1mL ADN, ADN/ARN,
rod ARN, mARN
AutoPure LS Centrifugation 16 96/jour Jusqu’à 10mL ADN
QIACube Filtration + 12 65min jusqu’à 200µL ADN, ADN et ARN
centrifugation viral, plasmid 51
51

52. Dedicated Workstations For NA extraction 4/4
QIAsymphony 300 LBC/16hrs; 1-96;
Qiagen Magnetic, Magnetic rod 2hrs up to 1ml DNA, RNA qiagen.com/
SP, RGQ 1-72
LBC, Urine Vials,
QIAEnsemble
Magnetic, Magnetic rod 3000 LBC 16hrs PreservCyt, surepath DNA qiagen.com/
SP
Vials, ETU tubes
Roche Diagnostics MagNAPure 2.0/96 Magnetic, Magtration; 32/96 2hr/1hr 20 à 300µL DNA, RNA, DNA/ roche-applied-science.com
PCR prep RNA, mRNA
MagNAPure Compact System Magtration; 8
Magnetic, 20min 100µL à 1mL
cartridge
AmpliPrep Magnetic,PCR prep, CE 1-72 24/1hr 100 à 750µL DNA, RNAv
IVD-D, continuous flow
Thermo LabSystems KingFisher Magnetic,Magnetic rod 2x12 24/17min, 29min, 20 à 200µL DNA, RNA, mRNA, thermo.com/
41min proteins, cells
KingFisher ML/96 15-96 15/14min, 36min, 10µL à 1mL
43min
52
52

53. A precursor in dedicated workstation
The ABI 340 extractor
A closed system for the process of 8 samples / 4 hrs.
53
53

54. From bench to automation
Phenol Chloroform extraction & Alcohol precipitation made automatic
54
54

55. Automated Platforms 1
Centrifugation Based
55
55

56. AutoGen Centrifugation based Workstations
AutoGen extractors
AutoGen 960-965 (4x96 samples in 4hrs)
AutoGenFlex Star Gene Prep
56
56

57. Autogen - 960-965 Extractors
1. Processing Method: • Single phase organic extraction
2. Protocols: DNA extraction from whole blood, blood spots, human
tissues and cells (including ES cells), plasmids, BACs, microbes and
plants.
3. Sample Volume:• Up to 1.8 mls per well
4. Maximum Sample Throughput:• 384 (96 deep well plate x 4) per
run
5. Processing Time:• Four hours for 4 plates
57
57

58. AutoGen - AutoGenFlex Star
An AutoGen and QIAGEN Collaboration
Removable sample
and DNA storage
racks
Heater to allow for
efficient digesting of
the sample and drying
of purified DNA
-4-position swing type buckets
-Variable speed and duration settings
-HEPA-Filter for biohazard reduction
High precision gripper-type robot
hand
Transports sample tubes to and
from workstation, dispensing
station and centrifuge
AutoGenFlex is completely
automated and easy to use, with no
manual inter vention required.
Isolate up to 80 x 10ml whole blood
samples per day
Qiagen's proven FlexiGene chemistry
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58

59. AutoGen - Gene PREP
On-Board Centrifuge and XYZ Robot
Heating Shaker
Optional Tube Unit Stacker
The GENE PREP is the latest fully
automated DNA extraction system
from AutoGen. •Automates entire extraction process including on board digestion.
•Capable of extraction from multiple sample types.
•Low cost per prep using AutoGen’s reliable solution phase extraction
chemistry.
48 samples in 2 hours (faster time to result).
Direct application of extracted nucleic acid to your •Higher throughput based on new design which shortens processing time.
next procedure (saves labor and time). •Produces high quality and high yields of DNA.
•Optional tube stacker allows for unattended processing of up to 192
samples.
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60. Gentra/Qiagen AUTOPURE LS
Complete automation of process for large volume samples
60
60

61. Genomic ExtraGene Platform
ExtraGene is a multi-protocol platform based on
centrifugation.
ExtraGene process up to 48 samples in 20 ml tubes.
The running time depends on the protocol.
Up to two improved centrifuges, with a
mechanized hatch on the top
The robotic arm moves a special tool for
tubes cap opening, closing, tube handling,
nozzles for syringe pumps and precision
peristaltic pumps
On the front side, a drawer supports tubes
racks, tips racks and liquid and solid waste
collector
ExtraGene is contained in a fume-hod
(optional)
Generic reagents = Much more extractions
with your budget
Full automation = Much more extractions
with your work force
Separation by centrifugation = Higher yeld
and lower cost (less than 1.5€ / extraction)
genomics-tools.com
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61

62. Automated Platforms 2
Filtration Based
62
62

63. Qiagen Extraction Platforms
BioRobot 3000
BioRobot 9604
63
63

64. Filtration based platforms
The BioRobot 8000
The BRmdx
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64

65. Filtration based platform on a centrifuge
The QIAcube process QIAGEN spin columns,
enabling seamless integration of automated,
low-throughput sample prep into your laboratory
workflow. No change of purification chemistry is
required, assuring fast startup and immediate
results. All steps in the purification procedure
are fully automated – including sample lysis –
and up to 12 samples can be processed per
run.
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65

66. FujiFilm QuickGene 810
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67. Complete Automation with the QuickGene HT
67
67

68. Beckman Coulter Inc.Vidiera NsP
Primary tubes on racks
NA extraction by filtration
Qiagen chemistry
Quantification by OD
Normalization
PCR setup
Archieving
68
68

69. Automated Platforms 3
Magnetic Beads Based
69
69

70. Biomerieux Nuclisens MiniMag
12 to 24 sample batch
70
70

71. Biomerieux Nuclisens EasyMag
“High throughput” nucleic acid automated extraction with minimum hands-
on-time and a very fast turnaround time (40 minutes for 24 extractions).
71
71

72. PSS, Japan
a precursor in magnetic extraction automation
Magtration Technology
72
72

73. PSS Workstations-1
PSS SX-6G PSS SX-8G
73
73

74. PSS Workstations-2
74
74

75. PSS Workstations-new concept
Pre-packed reagents for magnetic extraction procedures
75
75

76. A key evolution for NA extraction
Instrumentation: predisposable reagents
76
76

77. PSS Workstations-new concept
Individual barcoding for reaction cartridges
CCD camera procedure monitoring
77
77

78. PSS Workstations-new concept
Large volume magnetic extraction- PSS-8LX The new Magtration System BA-X
0.2 to 3 ml sample
volume handling
1 to 10 ml sample volume
50 min run
pre-filled reaction cartridge
78
78

79. Beckman Coulter Inc. SPRI-TE
Automated processing of 1-10 samples
in 30 minutes
✓ High-performance SPRI magnetic bead
technology
✓ Pre-packaged, ready-to-use reagents and
labware
✓ Simple to operate
✓ Method card driven protocols
✓ UV decontamination
79
79

80. Nordiag Magnetic Extraction
The affordable NorDiag Arrow
instrument makes automated
sample preparation possible for
all laboratories. The new
innovative tip pump assembly
will allows for a wide range of
sample volumes for extraction.
The tip pump assembly is a
closed system, reducing the
possibility for contamination.
Touch screen operation which
makes the instrument very user
friendly.
The NorDiag Bullet, 8 tip
config.
96 samples from primary tube to
PCR*/SDA** setup in less than The BUGS’n BEADS technology is
1.5 hours, using the NorDiag based on an initial adsorption of
Bullet instrument bacteria/cells to magnetic particles,
followed by a magnetic separation
where the sample material is
removed. This step removes most of
the inhibitors that are often found in
samples. The next step is lysis of cells
and the release of nucleic acids,
followed by immobilisation of nucleic
acids to magnetic particles.
80
80

81. Qiagen Magnetic Extraction
EZ1 Advanced EZ1 Advanced XL
BioRobot M48 workstation BioSprint 96 Workstation QIAsymphony Workstation
81
81

82. Roche Magnetic Extraction
COBAS® AmpliPrep-System
82

83. 2010 Evolution in Roche Automated Platforms
The Magna Pure 96 System
Time from sample to results: 2 hrs Speed - 96 sample extraction in <1hr
Easy loading
Prefilled reagent trays
Fewer disposables
Run surveillance
Connectivity
83
83

84. Magnetic rods
Qiagen
BioSprint
84
84

85. Promega Maxwell 16 System
Magnetic rods
16 sample batch
16 samples/30 min
85
85

86. Chemagic by Chemagen
96 rods for <200 µl samples in 96 well plates
4000 samples/day (200µl/6µg)
12 rods for <10 ml samples in 15 or 50 ml tubes
12 samples/40 min (7ml/210µg)
Distributed in the US by Parallabs
86
86

87. Chemagic by Chemagen
1 – 12 samples per run
๏ integrated buffer dispensing
๏ revolutionary resuspension technology
๏ standard plastic devices
๏ chemagic Kits and protocols for a huge variety of
sample materials
๏ quantitative yield, highest purity
๏ sample sizes up to 600 µl
๏ bar code reading available
87
87

88. NA extraction automation options
✓Microtiter Plate based Procedures
✓Generic Robotic Instrumentation
✓Dedicated Robotic Instrumentation
✓In Vitro Diagnostic Robotic Instrumentation
88
88

89. Abbott Molecular solutions
m2000sp m2000rt
RealTime RealTime RealTime VZV PCR
HIV-1 HBV mS9 Kit
RealTime RealTime Parvo B19 EBV PCR
HCV High Risk PCR Kit Kit
HPV
RealTime HSV 1/2 M.
HCV RealTime PCR Kit tuberculosi
Genotype CT/NG s
II CMV PCR
RealTime Kit SARS RT-
CT PCR Kit
89
89

90. Abbott Molecular solutions
Abbott M24sp (Freedom EVO Clinical 75)
1 to 24 sample batch processing
NA extraction, Open Mode
No PCR setup
CE-marked
90
90

91. Biomerieux NucliSens EasyMag Platform
“High throughput” nucleic acid automated extraction with
minimum hands-on-time and a very fast turnaround time (40
minutes for 24 extractions).
91
91

92. The Biomerieux Nuclisens EasyMag On Site
92
92

93. Qiagen MDx, CE-IVD Instrument
93
93

94. Qiagen platforms: New Generation
The QIAsymphony SP
•from 1 to 96 samples in batches of 24
•volumes up to 1 ml
•Magnetic Rod separation
•in-process sample loading
•bar code reading
•expandable
2008 2010 2011? 94
94

95. Qiagen QIAsymphony RGQ
The QIAsymphony RGQ:
from DNA extraction with the QSsp, to
PCR setup with the QAas and PCR with
the Rotor-Gene Q (1 to 72 samples per run)
HIV, HCV, HBV and CMV, EBV, HSV, VZV,
BKV1 Artus RealTime PCR kits available for
the Rotor-Gene Q. More to come.
95
95

96. Qiagen platforms: Next Generation
The QIAensemble
96
96

97. Roche Cobas AmpliPrep
Cobas Ampliprep + 2xTaqMan 48
97
97

98. Roche WNV TaqScreen platform
COBAS AmpliPrep COBAS TaqMan
Hamilton Pipettor
Pool Size = 6
98
98

99. Roche Cobas S201 System
99
99

100. Roche Cobas p630 System
The cobas p 630 Instrument
provides fully automated pre-
analytical solution for primary
tube handling.
It can be used in conjunction
with the COBAS® AmpliPrep/
TaqMan® System and can
support up to three COBAS®
AmpliPrep Instruments.
100
100

101. Roche Cobas 4800 System
Flexibility to meet different needs
• Adapts to the needs and throughput of each lab
• Can run up to 376 samples in a day but is still efficient for
labs with less intensive testing needs
Streamlined workflow
• Nucleic acid extraction from multiple types of specimens is fully automated,
with the fewest manual steps
• Swab and urine samples in cobas PCR Media in a
single run
• Multiple liquid cytology media types in a single run
• No manual preprocessing
• Cobas PCR Media tubes and liquid cytology vials can be loaded directly into
the system
• Load-and-Go reagents are already bar-coded and ready to use, requiring no
additional mixing
• Bidirectional connectivity to the LIS enables the automated reporting of results
101

102. Roche Cobas S401 System
102
102

103. Siemens Medical Solutions Diagnostics
Siemens
Competition
Primary tube sampling
Up to 96 samples per run
Proprietary extraction
technology using mag. particles
103
103

104. From sample Prep Automation to ? with the BD Viper
The BD Viper System is a patented
approach to laboratory automation
using an industrial class of robotics
known as Selective Compliance Assembly
Robot Arm (SCARA).
This classification indicates that the
robot is multi-jointed (4-axis), similar to
the human arm.
The BD Viper Sample Processor
automates the sample handling
associated with high volume amplified
molecular testing of Chlamydia
trachomatis and Neisseria gonorrhoeae
performed on the BD ProbeTec ET
System.
More than 500 samples for CT and GC
can be processed per 8-hour shift.
104
104

105. The BD Viper with XTR Technology
Throughput
Up to 736 CT/CG results in 8.5 hour shift
Time to First Results
I wish 3 hours 15 minutes
I could Amplification and Detection Technology
Strand displacement amplification with real-time
see the XTR amplification and detection
Contamination Prevention
Closed solid barrier amplification system
Specimen Collection Type
Vaginal & endocervical swabs, urethral swabs,
urine, liquid-base cytology (LBC). All collection
devices utilize pierceable caps during processing
On-Board Sample Capacity
96 samples including controls per run
Patient Sample Identification Location
A membrane switch pad records the location of the
tube. Capacity to read multiple barcode
configurations: 2 of 5, code 39, code 128, codabar
Sample Dispensing System
6-station pipettor with liquid-level sensing
Sample Volume
• On-board sample extraction utilizing proprietary • Zero reagent preparation – Ready-to-use reagents 800 µL
BD Fox™ chemistry • Automated sample cap piercing
• Walk-away platform with "load and go" processing • Liquid-level sensing of samples and reagents
• Automation using SCARA robotic technology • Patient sample location identification
• Multiple sample types – Liquid-based cytology,
male urethral swabs, endocervical swabs, vaginal
swabs, and urines
105
105

106. Gen-Probe DTS 800 Systems
DTS: Simplified Sample Processing
The DTS family of systems uses an exclusive penetrable cap to
minimize contamination and results in safer, more convenient
sample removal.
The design also reduces the time and effort required to remove
the cap and obtain swab samples from the tube.
DTS simplifies sample transport, minimizes handling and greatly
reduces laboratory cross-contamination.･In addition, bar code DTS® 800 Systems
technology simplifies the preparation of worklists. Generates up to 800 results in 8 hours
Includes the primary equipment and ancillary
Sample Preparation System items to process APTIMA assays
The Ten-Tube Unit (TTU) design
106
106

107. The Very 1st Fully Integrated MDx Platform
Complete automation of extraction, amplification and detection
one operator can test 500 samples with the APTIMA Combo 2 assay in approximately 8.5 hours, or 1,000
samples in approximately 13.5 hours. In addition, one operator can run two TIGRIS systems
simultaneously.
APTIMA Combo 2 assay for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (GC)
Procleix HIV-1/HCV Assay for simultaneously detection of HIV-1 and HCV
107
107

108. Gen-Probe - TIGRIS
108
108

109. Gen-Probe® Panther New System
1 the PANTHER system can continue running
unattended, processing 500 samples in 12
hours.
2 Primary tube sampling with random access
loading for maximum flexibility and productivity.
3 Multiple assays can be run from a single
sample, and operators have continuous access
to samples, reagents and consumables.
4 A customer-driven design, intuitive software for
ease of use, and remote diagnostics
capabilities.
5 Extensive process controls that include positive
sample identification, liquid level detection,
reagent dispense verification, and radio
frequency identification (RFID) tags on fluid
containers.
Random access
Optimized workflow and throughput
STAT test prioritization
Multiple tests from a single sample
In Europe, the PANTHER system can initially be used to perform Gen-Probe's nucleic
acid tests for Chlamydia and gonorrhea. These include the APTIMA Combo 2(R),
APTIMA(R) CT (Chlamydia) and APTIMA GC (gonorrhea) assays.
109
109

110. Fully Automated MDX Platforms EVolution
IVD industry Challengers:
Cepheid GeneXpert -> Cepheid Infinity
BD MAX System
Icubate, Idaho, Iquum, GeneFluidics
Upcoming or in Dev.
Hitachi High-Tech Open Platform
BCI UniCel DxN
110
110

111. What Future for NA Extraction in MDx?
✓Automation
✓Magnetic beads = most popular
✓Pre-dispensed reagents
✓Size of batch determination not final (6, 8, 24, 32, 48, 72, 96)
✓96 sample batch mainly found on “generic” robotics
✓Semi-random and random access platforms start to be popular
✓Evolution through complete integration of NA extraction + PCR + Detection
✓More questions unsolved about sample preservation
111
111

112. Archiving NA samples
112
112

113. DNA Storage
113
113

114. Storage of NA Samples at -80°C
114
114

115. Freedom EVO / REMP SSS Factory
The Freedom EVO® and
the REMP (Small Size
Store™ (SSS) together
enable the storage,
retrieval and
reformatting of
compounds and DNA
samples.
115

116. Biophile ultra-low temperature automated
Biophile (part of TekCel)
a combination of freezer, automation and information
technologies.
BIOPHILE, Inc., is the first to market -80C ultra-low temperature automated
storage and retrieval systems. 116
116

117. High Throughput Clinical Sample Storage
ARUP implemented the original Automated Transport and
Sorting System built by MDS AutoLab of Toronto, Ontario on
November 17, 1998. This system served ARUP's needs
extremely well for more than five years, reaching a level of
18,000-22,000 new patient specimens per day and a total
volume of tubes transported and sorted each day of 27,000
to 32,000.
117
117

118. Sample Storage at room Temperature
DNA storage at room temperature instead of -20 or -80°C
๏Biomatrica, San Diego, CA
SampleMatrix is a dissolvable compound that stabilizes DNA at room
temperature
SampleMatrix forms a protective seal around the sample as it dries,
effectively ‘shrink-wrapping’ the DNA.
๏Genevault, Carlsbad, CA
FTA paper on plate: GenPlates or GenTegra: Inorganic matrix
Outsourcing sample storage
๏Imagene, Bordeaux/Paris, France
Encapsulating DNA, RNA or proteins in an aluminium-capsule under zero
atmosphere
118
118

119. Sample Storage at room Temperature
Biomatrica
Preserve and store DNA at room
temperature
Biomatrica has developed SampleMatrix, to stabilize and
protect biological materials at room temperature without
degradation. The SampleMatrix technology was designed by
combining extremophile biology that enables long-term
survival of organisms in extremely dry environments with
synthetic chemistry.
Samples are preserved through formation of a protective
thermo-stable barrier during a simple air-drying process.
Samples are protected from degradation from heat and UV
light.
DNAstable enables you to store plasmid and genomic DNA
samples at room temperature. When you are ready to use
your samples, 100% of the DNA can be recovered in 10
minutes or less and then used directly in downstream
applications. By exploiting these unique characteristics,
Biomatrica products enable researchers to stabilize
biological samples for long periods of time.
119
119

120. Sample Storage at room Temperature
GenVault
GenPlates’ proprietary format features FTA paper elements
built into each wells for transport and long term storage of
biosamples at ambient temperature, in the dry-state.
Upon application of 10ul of biosample directly to the 6mm
disc of FTA paper, cells lyse and release DNA. The DNA
becomes entwined in the fibrous network of the cellulose
matrix and is dried completely at ambient temperature.
When DNA is required for downstream analysis, high-quality
double-stranded DNA from FTA paper can be reliably
recovered using the GenSolve chemistry.
Samples stabilized and dried down with GenTegra and
stored for over 12 months at ambient laboratory temperature
(21°C to 25°C) perform identically to DNA stored at -20°C in
all downstream applications. Long-term accelerated stability
studies of DNA at elevated temperature (76°C) indicate that
DNA stabilized with GenTegra will retain its molecular
integrity for decades at ambient laboratory temperatures.
Ensuring the dryness of DNA samples prior to storage is an
important factor in preserving DNA integrity in ambient
temperature storage conditions.
120
120

121. Sample Storage at room Temperature
Imagene
Room temperature DNA preservation
This technology relies in a DNAshell capsule which utilizes
encapsulation under a controlled anhydrous and anoxic
atmosphere of preliminarily desicated and purified DNA, into
small sealed, corrosion-proof metallic capsules. This process
prevents DNA from being altered by any degradation factors
(water, oxygen, light).
Completely anhydrous, anoxic and perfectly airtight, the
DNAshell capsules provides decisive advantages compared
to freezing, enhancing stability, safety, transport and
distribution of DNA. It also lowers operation and
maintenance costs. In addition, the laser-marked Data Matrix
codes on the capsules enable easy traceability and inventory
of each DNA sample.
Ensuring the dryness of DNA samples prior to storage is an
important factor in preserving DNA integrity in ambient
temperature storage conditions.
DNA can therefore be preserved at room
temperature for unlimited periods.
121
121

122. Robotics Archive or Personal Archive
GenVault’s DeskTop Archive is the
smallest in the series of room-
temperature biosample archiving
systems available from GenVault.
The “desktop” size system allows
one to test-drive the concept of
biosample storage at
roomtemperature at low cost and
low risk. Up to 1200 biosamples.
GenVault’s Dynamic Archive Biomatrica's dry storage cabinets
is an automated system for employ passive control devices to
archiving and retrieving ensure a stable humidity environment
GenPlates at room and are useful for laboratories where
temperature. The system moisture levels or fluctuation is an
utilizes a robot to access the issue. The cabinets can maintain
shuttles that contain trays on relative humidity at <40% for the
which GenPlates are housed. optimal room temperature storage of
Up to 31,500 geneplates. samples stabilized in Biomatrica’s
products. Dry storage cabinets
require no energy input and help your
lab save money and "go green."
122
122

123. Sample Management Softwares
Greiner
http://www.greinerbioone.com/
Central Labo Europe
http://www.central-labo-europe.com/
And also: Genvault, GenoLogics...etc. Biomatrica
http://www.biomatrica.com/
123
123

124. Sample Management Software: LabCollector
http://www.labcollector.com/features.php
124
124

125. Time for Questions
Or Time for Lunch?
125
125