New EZ-2 4.0 series evaporators

Genevac systems facilitate a wide range of DNA sample preparation and processing activities, ranging from straightforward small volume concentration of DNA pellets prior to PCR, through to high throughput processing of many purified DNA or oligonucleotide samples.  Where a dry sample of DNA is required, concentration to dryness in a centrifugal evaporator is the ideal method, presenting the DNA as a dry film in the tube of plate well.  Freeze drying of DNA samples is not recommended because the resultant DNA powder is easily blown about or moved by electrostatic charges.

DNA Concentration
Concentration of DNA from samples containing water or a mixture of water and alcohols is a simple process, which for most researchers requires the miVac DNA concentrator.  Concentration to remove a few hundred microlitres of alcohol from a DNA pellet takes approximately 10 minutes, or less.   This is one of the simplest applications of a Genevac system – in a miVac DNA concentrator, set the temperature, select the method for alcohols or water, and press start. 

Oligonucleotides & DNA Purification
The manufacture of oligonucleotides commonly has two steps where evaporation is required, evaporation of ammonia solutions following synthesis, and then subsequent evaporation of purified samples.   Protection of oligonucleotides from damage during drying is critical, especially where a tag or label has been attached to the DNA.  Two studies by Genevac users show that this can safely be achieved in a Genevac evaporator: 

• Comparing Evaporators for Drying Oligonucleotides - Catherine McKeen, Eurogentec SA A Rapid,
• Safe Technique for Drying Oligonucleotides - Dr Tim Watts, Wellcome Trust Human Genome Centre

Samples containing ammonia solution should be evaporated using Dri-Pure® (500g) to prevent sample loss and cross contamination.  When ammonia solution is subjected to a vacuum the ammonia degasses.  Degassing must be controlled or the resultant foam overflows the tube causing sample loss and cross contamination.  In the same way that you control degassing of a bottle of soda by opening it slowly, gentle application of vacuum is required, this coupled with the high rotors speeds of Dri-Pure keep every sample where it should be.  The latest versions of the EZ-2 come with such a method pre-programmed, and the method is simple to program on the HT-Series.
  
Where samples have been purified by reverse phase HPLC samples are presented in a mixture of water and methanol or acetonitrile.  The methods laid out in the Post Purification Sample Handling section should be followed.  The imbalance tolerance built into Genevac systems gives researchers confidence that eluates at different ends of the gradient will still dry down, and if there is too much imbalance the system will automatically shut down and tell the user.

Preparation of DNA Microarrays
For certain microarraying techniques concentrated samples of DNA are required.  To maintain such a concentration is difficult due to the volatile nature of the buffers holding the DNA, therefore samples should be made up to the required volume immediately prior to analysis.  A study by the Wellcome Trust Centre for Human Genetics in Oxford has shown that when an EZ-2 is used in this process the data quality returned by the system is significantly better, reducing the no call rate by 80% or more.

Quality and safety testing of food and beverages is an area of growing importance, be that for exporters, importers or government bodies, accurate analytical results are critical.  In common with many other applications, careful sample preparation is critical, especially when the analyte of interest is volatile.

Genevac systems find application in a number of areas, from testing constituents of beverages such as Cognac or gluten levels in Whisky, to pesticide analysis of fruit and vegetables, antibiotic residues in meats or vitamin levels in cereals.  The research and development of Functional Foods where high quality scientific research is carried out also benefits from the speed and quality of a Genevac system.  The EZ-2 or Rocket are commonly used, often in conjunction with SampleGenie™, because this delivers automation of sample transfer and gives unparalleled sample recovery and inter-test reproducibility with very low standard deviations, as the studies from the Laboratoire départemental d'analyses de la Drôme clearly establish.
Concentration technology in Genevac systems has been developed with leading analytical laboratories.  This, with key technologies like Dri-Pure, ensure that samples are concentrated safely and rapidly. SampleGenie™ is a great aid to concentration because the evaporator only evaporates the solvent contained in the large flask, and not the solvent in the vial.  The system detects the end point of evaporation being when the solvent level enters the vial and, once validated, the method will concentrate the sample to the required level.  If a precise volume is required in the vial, then the sample must be over concentrated and then made up to the desired level with pure solvent.

For protein analysis, e.g. for nutritional studies, the EZ-2 with HCI resistance option is ideal.  Protein digests often require digestion with 6N HCl which is highly corrosive to traditional evaporation systems, however, HCl resistant Genevac systems are designed to withstand daily use of concentrated HCl.  

The Rocket has also found an interest application in some of the worlds best restaurants, those that specialise in molecular cuisine, where it is used to concentrate sauces, juices and other natural flavours for the chef to use.  Details of how the Rocket is used can be found in a new book; Modernist Cuisine - the Art and Science of Cooking. The Studio Kitchen blog has some interesting receipe ideas, and some of these are also on Kitchen Theory. 

Useful Papers:

• An Improved Evaporative Sample Preparation Methodology for Determining Nitrofuran Antibiotic Residues in FoodstuffsKantonales Labor Zurich (KLZH), Zurich, Switzerland 
• Analysis for pesticides: Method development to increase the recovery of volatile compounds by using EZ-2, a new generation of centrifugal evaporatorEnvironmental Protection Agency of Tuscany (ARPAT), Florence,  Italy        
• Evaluation of the EZ-2 for Pesticide Analysis Laboratoire départemental d'analyses de la Drôme, Valence, France  
• Evaluation of SampleGenie™ for Environmental AnalysisLeochimica Laboratories, Zoppola, Italy 
• Evaluation of the Rocket and SampleGenie for Pesticide AnalysisLaboratoire départemental d'analyses de la Drôme, Valence, France

Natural products are researched for many reasons, from biodiversity preservation through flavour or fragrance research, to finding a cure for disease.  Whatever your reason, Genevac systems are used by researchers who are investigating the properties of the natural environment. 

For basic botanical and biological research, the miVac concentrator range are the mainstay for those extracting and purifying DNA or proteins for genome and proteome research.

Genevac systems are widely used by researchers seeking to extract functional molecules from natural sources.  A typical process would be to take a tissue (or micro-organism broth) sample and then perform a primary solvent extraction resulting in a large volume of crude extract.  The crude extract is often concentrated, for which the Rocket Evaporator is ideal, before being fractionated to isolate each component.  Fractionation may use a system such at the Sepbox, one, or two dimensional chromatography which produces many samples from one extract.  These samples need to be evaporated before storage or analysis and may require lyophilisation. Genevac HT series evaporators are used for this purpose. 

Useful Papers:

• High Throughput Natural Product Discovery - a paper by Stephen Pickrahn, IFF 
• Rapid Evaporation Processes to Enhance Natural Product Extraction - by Dr John Hester, Principal R&D Engineer – Natural Product Repository Supervisor, University of Mississippi

Protein concentration is an issue that the biological sciences have been dealing with for years. Many aspects of the study of proteins and exploitation of their therapeutic value requires concentration before and after processing. Each process step can range in scale from setting up a sitting drop of 100nl in microtitre plate based crystallization study to purification of the product from 10,000L fermentation. All of the currently available tools have their draw-backs whether it is from yield loss on membranes to dilution from capture columns.

One of the most commonly used techniques at all scales is membrane concentration. At the small scale this is usually based on concentrators that fit in a centrifuge or small pressure based filers both of which rely on dead-end filtration to push the supernatant through a membrane and retain the protein. The small scale methods used suffer badly from concentration polarization a phenomenon observed in membrane concentrations where the protein concentration at the membrane increases due to the removal of liquid, this is analogous to the cake formed in conventional filtration. In larger applications a cross flow membrane is commonly used where the pressure potential for filtration is provided by a tangential flow to the membrane surface.This cross flow helps to reduce the effect of concentration polarization by sweeping the membrane to clear the retained protein; however, due to constant solvent removal within the system there is still a polarization gradient setup at steady state. When a protein reaches ultra-high concentration at the membrane interface then gel formation is likely this can reduce the activity of the protein and, if the regime for re-suspension is not effective, lead to loss of yield. When we couple this to the non-specific binding effects of most membrane systems it makes this a technique that the industry is using for necessity rather than optimized performance.

One alternative for laboratory scale concentration is to use Centrifugal Vacuum concentration where the solvent is boiled from the supernatant under vacuum so that the temperature of boiling is below the denaturation threshold of the protein. This can dramatically improve both the yield after concentration and also the activity of the protein.This technique would not be used on the large scale but finds a niche in the small scale laboratory work, for example preparation of sample prior to screening against a panel of precipitants to determine the optimum conditions for crystallisation prior to XRD. For hanging and sitting drop experiments concentrations greater than 10 to 20 mg/ml are often required, this is very difficult to achieve in membrane based concentrators due to the limiting factors that have been highlighted above. Where the sample has been purified using reverse phase chromatography (typically peptides), leaving the sample in water and organic solvent, this cannot be concentrated using a membrane because the organic solvents damage the membrane. In a vacuum concentrator the sample remains in a micro-centrifuge tube and the volume remaining can be accurately determined by the time of concentration. One potential limitation of this technique is that the salt and buffer components of the solution will also be concentrated, hence this is ideally suited to be a polishing step that is used after a capture column or membrane concentration step and therefore allows the sample to reach the very high concentrations needed for crystallization. The most commonly used systems for these applications are the miVac range of biological concentrators or the EZ-2.

Peptide Synthesis
Peptide synthesis is normally performed using solid phase supported stub with the sequential addition of protected amino acids. Between each addition step, the protected terminal needs to be deprotected so that the next acid can react and join.  When the desired peptide sequence is complete, it is cleaved from the solid support, usually using a strong acid such as TFA.  The peptide in cleavage cocktail is then dried and purified. Following purification the peptide can be fast lyophilised in a Genevac HT series evaporator or dried using a traditional freeze drier.

Useful Papers:

• Better Analysis of Insulin Analogues - Dr Catrin Goebel, Australian Sports Drug Testing Lab Examining the effects of sample preparation on data qualityControlled
• Concentration for MALDI-TOF MS - Steve Knight, GenevacLooking at the effects of sample preparation on MALDI targets in cancer research

Contact us if you are interested in any of the above papers

Working with oils, fuels and fats can be difficult.  Whether extracting & recovering additives, evaluating lipid content of foods, characterising reclaimed cooking oils before reprocessing or conversion to biofuels, or investigating drillling test samples - oils can be difficult to dry in turn leading to problems with the analytical instrumentation.   

Extraction and separation techniques, be they basic dialysis, Dionex ASE or Counter Current Chromatography, all normally present the sample in a solution of organic solvent.  The sample typcially must be dried or concentrated before analysis.  Drying all the solvent can be difficult without using very high temperatures or other harsh regimens which may lead to loss of more volatile fractions within the oil.  For many such applications, researchers have found that the EZ-2 and the Rocket Evaporator have delivered significant enhancement to their recovery and results.  When working with larger solvent volumes, additional benefit is gained by way of  productivity savings when used with SampleGenie which enables large volumes to be concentrated or dried directly into a small vial.