Lab Equipment

High-Field & High Definition 700MHz NMR with four channel Cryoprobe

1. Magnet and Shim system

Actively shielded superconducting magnet system with a min. field strength of 16,4 Tesla and with a room temperature bore of min. 54 mm.
Advanced room temperature shim system for optimal magnet homogeneity as required for 1D-, 2D-und 3D experiments without sample tube rotation.
Nitrogen liquefier (Nitrogen hold time > 300 days)
Room temperature shim system with 36 gradients, low power technology with both manual and software controlled power supply with automatic shim function of Z- and Off axis gradients 1H and 2H (lock signal)

2. Lock System

Frequency variable 2H lock system with quadrature detection and digital filters for highest filter performance, for highest lock sensitivity and for software/user controllable optimisation of lock parameters.

3. Variable Temperature Control

Software controlled variable temperature unit, adjustable in 0.1 °C steps for the control of a temperature range from – 150 °C to + 200 °C, stability: 0.01 °C.
Cooling accessory to provide pre cooled air for operation temperatures from -5 °C to ambient & – Variable temperature range is from 0°C to +80°C

4. Cryo Accessory

Cryoplatform consisting of cryo-cooling unit with integrated nitrogen liquefier to extend the nitrogen hold time of the magnet up to one year.
Air-cooled Helium compressor
Cryogenic triple resonance probe: For the analysis of biological samples and mixtures in limited concentration the following cryogenic probe is required to achieve highest sensitivity:

Inverse triple resonance high resolution cryogenic probe fitted with an actively shielded single axis Z-gradient for 5mm sample diameters for highest 1H and 13C sensitivity, short pulse widths and highest 2H sensitivity for maximal lock stability is required. This probe needs to be capable of usual inverse triple resonance experiments and also of 13C observation and decoupling experiments. Highest possible 13C sensitivity is essential for the structure determination of natural products and metabolites. The probe is fitted with an actively shielded single axis Z-gradient and with a 2H lock channel.
Automatic tuning and matching capabilities for all tree channels are required.

The high-field, high-definition, 700MHz NMR at the Dept of Pharmacy, U of Patras, acquired through FP7-REGPOT “SEEDRUG” project (Nr. 285950, www.seedrug.upatras.gr)

Fast Protein Liquid Chromatography (FPLC)/ High Pressure Liquid Chromatography (HPLC)

The laboratory’s equipment includes four automated FPLC systems from ÄKTA GE Healthcare Life Sciences, an ÄKTA Purifier 900, an ÄKTA Pure 25M, an ÄKTA GO and an ÄKTA START, which is placed in the new COLD ROOM facility of the lab.

ÄKTA pure 25M can be used for High Pressure Liquid Chromatography and is designed for a broad range of applications and purification tasks in a multi-user environment.

Features:

Purification systems appropriate for proteins, peptides and nucleic acids, from microgram to gram scale.
Dual pump systems, allowing to run two different solutions simultaneously.
The flow rate of AKTA’s is between 0,01ml/min and 10ml/min, whereas the flow rate of AKTA pure25M can reach up to 25ml/min.
Variable multiwavelength UV and Visible spectrum monitor from 190 to 700nm.
Monitoring up to three wavelengths at the same time.
Online pH and conductivity detectors are adjusted to the systems
Automated fraction collectors.
Different injection volume loops are available from 10μl-loops to 5ml super-loops.
Wide range of columns (Superdex 200 10/300GL, Superose 12 10/300GL, Mono Q 5/50GL, Mono S 5/50GL etc.) for different types of liquid chromatography are compatible to both systems and available from our laboratory

Applications:

Chromatographic Analysis of biomolecules, proteins polypeptides, peptides, polymers and biosurfactants.
Small scale preparative purification (flow rate of AKTA pure25M can up to 25ml/min and pressure limit 20MPa).

FUNDING/INSTALLATION (YEAR)

ÄKTA Purifier 900 (Bodossaki Foundatiuon & Research Committee, UPAT, 2013)

ÄKTA pure 25M: (FP7-SEEDRUG/285950, 2015)

ÄKTA GO & ÄKTA START (Region of Western Greece, Support of National RIs/Κ.Ε. 2020ΕΠ00100001, 2021)

SEC-MALS

SEC-MALS is a powerful biophysical method, which combines multi-angle light scattering with size-exclusion chromatography. It is capable to characterize macromolelcular properties like:

Molar mass
Size
Conformation
Conjugation ratio

In SEC-MALS, the SEC column serves to separate molecules by hydrodynamic volume. Retention time is not used to determine molecular weight. Instead, each HPLC/FPLC fraction exiting the SEC column pass through a MALS detector and the solution (and the biomolecules included) are probed by a laser beam.

The WyattQELS module is embedded within a DAWN 8 online multi-angle light scattering (MALS) instrument in order to determine high-resolution particle size distributions, in conjunction with a separation technique size exclusion chromatography- (NEXERA series/SHIMADZU Ultra High-Performance Liquid Chromatograph). In SEC-MALS, the SEC column serves to separate molecules by hydrodynamic volume; retention time is not used to determine molecular weight. After exiting the column of the UHPLC the molecules pass through a MALS detector and are probed by a laser beam. Then, the MALS signals, along with UV absorbance and differential refractive index (dRI) signals, are analyzed to quantify the analyte’s physical properties.

Features:

SEC: UV-VIS Detector: SPD-40/40V, Photodiode Array (PDA) Detector: SPD-M40Solvent, Delivery Pump: LC-40D series, Autosampler: SIL-40C series, Degassing Unit: DGU-405, Column Oven: CTO-40C series
WyattQELS: Size Range (Rh), flow mode 0.5 nm to 300 nm
Molar Mass Range 200 Da to 100 MDa
Molecular Size Range (Rg) ≈ 10 to 300 nm, up to 600 nm with shape-specific models
The DAWN is accompanied by ASTRA software package available for analysis of online multi-angle and dynamic light scattering.

Applications:

Absolute characterization of the molar mass and size of proteins, conjugates, macromolecules and nanoparticles in solution
Determination of distributions of mass, size, and composition independent of column calibration by reference standards.

FUNDING/INSTALLATION (YEAR)

Region of Western Greece, Support of National RIs/Κ.Ε. 2020ΕΠ00100001, 2021

Isothermal Titration Calorimetry (ITC)

Our lab applies thermodynamic techniques. In this direction, we are equipped with Malvern MicroCal Peaq Isothermal Titration Calorimetry (ITC). It allows direct, label-free in solution measurement of binding affinity and thermodynamics in a single experiment, enabling the accurate determination of binding constants (KD), reaction stoichiometry (n), enthalpy (ΔH) and entropy (ΔS)

Features:

Batch evaluation of large data sets, automated assessment of data quality
Automated washing with detergent of the sample cell and titration syringe assists in producing high quality reproducible data
Sensitive enough to investigate biomolecular interaction using as little as 10 μg protein
Specification: Sample volume: 280μL, Sample cell size: 200μL, Injection syringe volume: 40μL, Injection volume precision: <1% @ 2μL
Throughput: 8-12 per 8h day
Noise: 0.15 ncal/s
Temperature Range: 2 to 80
Response time: 8s
MicroCal PEAQ-ITC analysis software

Open multiple experiments in a single session
Automated fitting models (One-Site, Two-Site, Sequential, Competitive, Enzyme Kinetics, Dissociation)
Automated assessment of data quality: Good quality data – Binding, Good quality data – No binding, Poor quality data – Check data

Applications:

Characterizing biomolecular interactions, to:

Confirm binding and activity
Determine stoichiometry and thermodynamic parameters
Study structure activity relationships

Studying the interaction of any two biomolecules including:

Proteins, nucleic acids, lipids, drugs (APIs).

FUNDING/INSTALLATION (YEAR)

Region of Western Greece, Support of National RIs/Κ.Ε. 2020ΕΠ00100001, 2021

Circular Dichroism Spectrometer (CD)

To determine protein’s secondary structure our lab is equipped with Chirascan qCD Circular Dichroism Spectrometer from Applied Photophysics.

Features:

The key performance capabilities of Chirascan qCD are listed below relate directly to accuracy, speed, sensitivity, and lower running costs:

Multi-point absolute CD calibration using an optical standard
Higher light throughput than all other CD spectrometers (which means better far-UV performance)
Temperature ramping capabilities (e.g. multiple T-ramp curves in one experiment)
Global thermodynamic analysis software
Chirascan qCD offers a high performance PMT detector (range 163nm – 950nm) with triple detector capability for CD, LD and ORD measurements without any physical reconfiguration to the system.
Automated shutter before the monochromator which not only prevents the sample from the access light (photo-bleaching effect) but also the optics from being degraded in a longer run, increasing the lifespan of the instrument
Single Cell Pelter Cell Holder. The Single Cell Peltier Temperature Controller features a wide operating temperature range of -20°C to 110°C, using a circulating chiller unit, excellent thermal contact and accommodates both rectangular and cylindrical cuvettes.
Better suited for high scattering samples: Chirascan’s detector is located very close to the sample cell which maximises light collection for highly scattering samples such as membrane proteins. If required, the detector’s position can be easily positioned further from the sample cell.

Applications:

Suitable for studying chiral molecules of all types
Determine the content of protein secondary structure
Thermal denaturation (temperature ramping) experiments provide information on stability, mechanisms of folding/unfolding including melting temperature (Tm).

FUNDING/INSTALLATION (YEAR)

H2020-MARISURF/635340 (2017)

Glove box

Some proteins/biomolecules due to their sensitivity in oxygen have special requirements in which we should work. To meet these special needs, we installed the PureLab HE 2GB Glovebox System. This system provides air-free (O2) and water-free (H2O) processing capabilities in a humidity controlled glove box. The presence of argon or nitrogen gas is a consistent condition needing managing and monitoring.

Features:

Glove box expansion features include additional antechambers for thru-flow work processes, cold storage freezers, process vacuum ovens and furnaces, and a full line of accessories.

Inert’s Gas Management Systems provide a < 1ppm O₂ and H₂O inert atmosphere required for a multitude of air-sensitive applications
Vacuum pump: 10 cfm (17 m³/h) rotary vane pump with oil mist eliminator and oil return kit
Integrated oxygen analyzer: Range 0-1000 ppm
Integrated moisture analyzer: Range 0-1000 ppm
Working gas: 99.995% N₂ or Ar, pressure: 55 psi (3.8 bar), gas flow rate: 250 l/min

Applications:

Protein production in controlled atmosphere to prevent unwanted chemical reactions such as oxidation and hydrolysis that is degrading a sample.
Other applications that require O₂-free atmosphere and low humidity environment.

FUNDING/INSTALLATION (YEAR)

National RIs “INSPIRED”/MIS-5002550 (2020)

ChemiDoc Imaging System

The ChemiDoc Imaging System provides fast, reliable, and sensitive imaging and documentation of gels and chemiluminescence western blots. This system is compatible with stain-free technology, chemiluminescence detection, and a wide range of gel stains such as ethidium bromide, SYPRO Ruby, Coomassie, and silver stains.

Features:

All-in-one flexible imaging: get precise, reproducible chemiluminescence and colorimetric blot detection, analysis, and gel documentation in a single system
Sample Trays and Smart Tray Technology: three sample trays available to cover diverse imaging applications. Smart Tray Technology automatically recognizes the application-specific trays and adjusts imaging parameters and software options accordingly
Stain-free protein normalization: stain-free imaging permits the normalization of bands to total protein in both in gels and blots; eliminate the need for housekeeping proteins
Ease of use: features include automatic selection of optimal light source by application, auto focus, auto exposure, and preview features ensure optimal images. Learn in minutes with on screen help
Component of the ChemiDoc Imaging System is the Trans-Blot Turbo System. An integrated five-step system to streamline your entire western blotting protocol:

Quality protein transfer in 3 min, transfer 1–4 mini gels or 1–2 midi gels in a single run
Higher transfer efficiency than other protein blotting methods
Faster and more efficient transfer than wet tank blotting

Applications:

Electrophoresis: protein, including stain-free, and nucleic acid gel imaging and documentation
Blotting: identify and quantify proteins, nucleic acids, protein-ligand interactions, etc. with chemiluminescence, or colorimetric detection
High quality images for reports/publications

FUNDING/INSTALLATION (YEAR)

National RIs “INSPIRED”/MIS-5002550 (2020)

COLD ROOM Facility 230 x 260 x 230(h) cm, temperature 4ο to 6οC.

FUNDING/INSTALLATION (YEAR)

Region of Western Greece, Support of National RIs/Κ.Ε. 2020ΕΠ00100001, 2021

NanoDrop Flurospectrometer

The Thermo Scientific NanoDrop™ 3300 Fluorospectrometer uses patented sample retention technology (surface tension) to make small volume fluorescent measurements. The uniquely clean optics of the retention system, combined with proprietary signal processing for the White LED applications, enables measurements across a wide range of wavelengths using sample volumes of 1–2 ul without cuvettes and costly filter changes.

The NanoDrop 3300 significantly lowers the mass detection limit more than an order of magnitude, compared to conventional fluorometers. With its advanced features and analysis options, the NanoDrop 3300 allows for a diverse range of measurements from routine RiboGreen, PicoGreen, and GFP to advanced nanoparticle and FRET analysis.

Features:

Minimum Sample Size: 1 µl
Fast measurement time of less than ten seconds
UV, Blue and White LED excitation, covering a broad wavelength range
Broad excitation range without the need to change filters or use of a monochromator
Ability to run diverse applications
Measures multiple fluorophores from a single sample
Tools to eliminate the guesswork
Provides both spectral and quantitative data

Applications:

Nucleic Acid, DNA, RNA and Protein Quantification
Binding interactions

FUNDING/INSTALLATION (YEAR)

National RIs “OMIC-ENGINE”/MIS-5002636 , Anticipated 2021

Sorvall LYNX 6000 Superspeed Centrifuge

To support high-performance separations our lab is equipped with five centrifuges:

Thermo Scientific™ Sorvall LYNX 6000 Superspeed Centrifuge
Thermo Scientific™ SL 40 Centrifuge Series
Beckman Coulter Allegra® X-15R
Thermo Scientific™ Fresco™ 21 Microcentrifuge
Thermo Scientific™ MicroCL 21R Microcentrifuge

Sorvall™ LYNX 6000 Superspeed Centrifuge features powerful technologies with breakthrough simplicity, from push-button security and application flexibility of Auto-Lock rotor exchange, to immediate rotor detection and programming of Auto-ID instant rotor identification, and the improved ergonomics and performance of carbon fiber rotors.

Features:

Capacity: up to 6L
Speed: 29,000 rpm
RCF: 100.605 x g
Temperature Range: -20°C to +40°C
Quickly and securely change rotors and applications in as little as 3 seconds with push-button Auto-Lock rotor exchange
Auto-ID instant rotor identification automatically loads rotor specifications into centrifuge parameters
19 optional rotors to find the ideal fit for your application supporting labware from microplates, to microtubes, to 1L bottles

Applications:

Centrifugation products are useful for functional proteomics, research involving protein-protein interaction studies, protein biochemical studies, large-scale protein folding, and 3-dimensional protein structure determinations

FUNDING/INSTALLATION (YEAR)

National RIs “OMIC-ENGINE”/MIS-5002636 (2019)

Shakers-Incubators

Infors HT Multitron Standard:

Tall enough for 5 L Erlenmeyer flasks
Can be set up as either a free-standing unit or benchtop unit, or in a three-tier stacking configuration

2x Infors HT Minitron

FUNDING/INSTALLATION (YEAR)

FP7-NEUROCYPRES/202088, 2009
FP7-SEEDRUG/285950, 2015,
National RIs “INSPIRED”/MIS-5002550

Benchtop Bio-reactor

Infors HT Minifors bio-reactor: This Bio-reactor is frequently used as a starting point for research and education using fermenters.
Minifors bio-reactor provide us pH control, feeding, sampling and software for data-logging.

Features:

All components integrated into a single housing
Heating block for temperature control
Removable, autoclavable pump heads (minimum of 2, maximum of 4)
Temperature: Up to 60°C, electrical heater block
Stirrer speed (Bacterial version): 100 – 1250 rpm
Stirrer speed (Cell version): 30 – 300 rpm
Vessel sizes: 2.5 L
Parameters: °C, rpm, pH, pO2 +AF, feed, optional additional parameters such as mass-flow, turbidity with optional analog input and outputs.

Applications:

Yeast and bacterial cultures

FUNDING/INSTALLATION (YEAR)

FP7-SEEDRUG/285950, 2015

PCR

BioRad T100™ Thermal Cycler:

The T100 thermal cycler is a small thermal cycler offering a comprehensive set of convenient features in a small footprint. This compact thermal cycler features an intuitive touch-screen user interface to make running PCR easy. Thermal gradient technology allows you to quickly optimize your reaction in a single run.

Applications:

Nucleic acid amplification (PCR)
Gene cloning and analysis
Gene expression analysis
Mutational analysis
Cycle sequencing

Lab Equipment

High-Field & High Definition 700MHz NMR with four channel Cryoprobe

1. Magnet and Shim system

Actively shielded superconducting magnet system with a min. field strength of 16,4 Tesla and with a room temperature bore of min. 54 mm.

Advanced room temperature shim system for optimal magnet homogeneity as required for 1D-, 2D-und 3D experiments without sample tube rotation.

Nitrogen liquefier (Nitrogen hold time > 300 days)

Room temperature shim system with 36 gradients, low power technology with both manual and software controlled power supply with automatic shim function of Z- and Off axis gradients 1H and 2H (lock signal)

2. Lock System

Frequency variable 2H lock system with quadrature detection and digital filters for highest filter performance, for highest lock sensitivity and for software/user controllable optimisation of lock parameters.

3. Variable Temperature Control

Software controlled variable temperature unit, adjustable in 0.1 °C steps for the control of a temperature range from – 150 °C to + 200 °C, stability: 0.01 °C.

Cooling accessory to provide pre cooled air for operation temperatures from -5 °C to ambient & – Variable temperature range is from 0°C to +80°C

4. Cryo Accessory

Cryoplatform consisting of cryo-cooling unit with integrated nitrogen liquefier to extend the nitrogen hold time of the magnet up to one year.

Air-cooled Helium compressor

Cryogenic triple resonance probe: For the analysis of biological samples and mixtures in limited concentration the following cryogenic probe is required to achieve highest sensitivity:

Automatic tuning and matching capabilities for all tree channels are required.

Fast Protein Liquid Chromatography (FPLC)/ High Pressure Liquid Chromatography (HPLC)

The laboratory’s equipment includes four automated FPLC systems from ÄKTA GE Healthcare Life Sciences, an ÄKTA Purifier 900, an ÄKTA Pure 25M, an ÄKTA GO and an ÄKTA START, which is placed in the new COLD ROOM facility of the lab.

ÄKTA pure 25M can be used for High Pressure Liquid Chromatography and is designed for a broad range of applications and purification tasks in a multi-user environment.

Features:

Purification systems appropriate for proteins, peptides and nucleic acids, from microgram to gram scale.

Dual pump systems, allowing to run two different solutions simultaneously.

The flow rate of AKTA’s is between 0,01ml/min and 10ml/min, whereas the flow rate of AKTA pure25M can reach up to 25ml/min.

Variable multiwavelength UV and Visible spectrum monitor from 190 to 700nm.

Monitoring up to three wavelengths at the same time.

Online pH and conductivity detectors are adjusted to the systems

Automated fraction collectors.

Different injection volume loops are available from 10μl-loops to 5ml super-loops.

Wide range of columns (Superdex 200 10/300GL, Superose 12 10/300GL, Mono Q 5/50GL, Mono S 5/50GL etc.) for different types of liquid chromatography are compatible to both systems and available from our laboratory

Applications:

Chromatographic Analysis of biomolecules, proteins polypeptides, peptides, polymers and biosurfactants.

Small scale preparative purification (flow rate of AKTA pure25M can up to 25ml/min and pressure limit 20MPa).

FUNDING/INSTALLATION (YEAR)

ÄKTA Purifier 900 (Bodossaki Foundatiuon & Research Committee, UPAT, 2013)

ÄKTA pure 25M: (FP7-SEEDRUG/285950, 2015)

ÄKTA GO & ÄKTA START (Region of Western Greece, Support of National RIs/Κ.Ε. 2020ΕΠ00100001, 2021)

SEC-MALS

SEC-MALS is a powerful biophysical method, which combines multi-angle light scattering with size-exclusion chromatography. It is capable to characterize macromolelcular properties like:

Molar mass

Size

Conformation

Conjugation ratio

Features:

SEC: UV-VIS Detector: SPD-40/40V, Photodiode Array (PDA) Detector: SPD-M40Solvent, Delivery Pump: LC-40D series, Autosampler: SIL-40C series, Degassing Unit: DGU-405, Column Oven: CTO-40C series

WyattQELS: Size Range (Rh), flow mode 0.5 nm to 300 nm

Molar Mass Range 200 Da to 100 MDa

Molecular Size Range (Rg) ≈ 10 to 300 nm, up to 600 nm with shape-specific models

The DAWN is accompanied by ASTRA software package available for analysis of online multi-angle and dynamic light scattering.

Applications:

Absolute characterization of the molar mass and size of proteins, conjugates, macromolecules and nanoparticles in solution

Determination of distributions of mass, size, and composition independent of column calibration by reference standards.

FUNDING/INSTALLATION (YEAR)

Region of Western Greece, Support of National RIs/Κ.Ε. 2020ΕΠ00100001, 2021

Isothermal Titration Calorimetry (ITC)

Features:

Batch evaluation of large data sets, automated assessment of data quality

Automated washing with detergent of the sample cell and titration syringe assists in producing high quality reproducible data

Sensitive enough to investigate biomolecular interaction using as little as 10 μg protein

Specification: Sample volume: 280μL, Sample cell size: 200μL, Injection syringe volume: 40μL, Injection volume precision: <1% @ 2μL

Throughput: 8-12 per 8h day

Noise: 0.15 ncal/s

Temperature Range: 2 to 80

Response time: 8s

MicroCal PEAQ-ITC analysis software

Applications:

Characterizing biomolecular interactions, to:

Confirm binding and activity

Determine stoichiometry and thermodynamic parameters

Study structure activity relationships

Studying the interaction of any two biomolecules including:

Proteins, nucleic acids, lipids, drugs (APIs).

FUNDING/INSTALLATION (YEAR)

Region of Western Greece, Support of National RIs/Κ.Ε. 2020ΕΠ00100001, 2021

Circular Dichroism Spectrometer (CD)

To determine protein’s secondary structure our lab is equipped with Chirascan qCD Circular Dichroism Spectrometer from Applied Photophysics.

Features:

The key performance capabilities of Chirascan qCD are listed below relate directly to accuracy, speed, sensitivity, and lower running costs:

Multi-point absolute CD calibration using an optical standard

Higher light throughput than all other CD spectrometers (which means better far-UV performance)

Inert’s Gas Management Systems provide a < 1ppm O₂ and H₂O inert atmosphere required for a multitude of air-sensitive applications

Vacuum pump: 10 cfm (17 m³/h) rotary vane pump with oil mist eliminator and oil return kit

Working gas: 99.995% N₂ or Ar, pressure: 55 psi (3.8 bar), gas flow rate: 250 l/min

Other applications that require O₂-free atmosphere and low humidity environment.