InnovaPrep’s contract # GS-07F-0747X

 InnovaPrep’s eleven pending and awarded patents apply to highly efficient collection, recovery, and concentration of biological particles from air, surfaces, and liquids.   InnovaPrep’s novel biological particle elution process underlies many of these patents and enables recovery of particles from filters, membranes, surfaces, and objects. The primary utility for these technologies is to greatly improve the way biological samples, especially those containing pathogenic organisms, are collected and prepared for analysis beyond the state-of-the-art.  Specifically, these technologies fill a critical void that currently severely limits application of the most advanced biological detection systems to the billions of microbiological tests performed annually.  This advance is 150 years overdue.

InnovaPrep recently underwent an independent performance review by Duns and Bradstreet as a requirement for becoming a government vendor with GSA. We submitted the names of 15 companies we have done business with in the past year, D&B then conducted a survey of their experiences doing business with us.

We were pleased to receive overwhelmingly positive ratings by our clients in all categories: reliability 94%, cost 91%, order accuracy 90%, timeliness 93%, quality 92%, business relations 95%, personnel 98%, customer support 95% and responsiveness 95%.

We sure appreciate our clients and see that though rated at the top of our industry, there is room for improvement and we certainly endeavor to continue to improve. Our goal is to be a world class provider of cutting edge solutions for sample collection, preparation and concentration that bridge the gap between the laboratory and real world environments.

~Ann

The Knowledge Foundation Sample Prep 2011 conference this year was even better than last year.  For one thing, it was in San Diego – but not that I didn’t enjoy Baltimore, too!  InnovaPrep was only one of many companies, Universities, and agencies that presented new developments this year.  Participants included Sandia, the University of California San Diego, Quidel,  Harvard Medical School, Boreal Genomics, Qiagen, Taigen, Merck Millipore, Boston University, Stratec, Iowa State University, Menon & Associates, the Canadian Food Inspection Service, Nanyang University of Singapore, FLIR Systems, Integrated Nano Technologies, Receptors LLC, NOAA, Claremont Bio Solutions, Deton Corp., USEPA, Hitatchi, Advanced Liquid Logic, and JSR Micro.  I also met with attendees from Fluigent, EcoLab, Pall Life Sciences, Bertin Technologies, bioMerieux and many others.

If you were there, or interested in a conversation about sample prep for rapid microbiological methods, we can get one started right here – just add your comments to the blog and we’ll post them!  I’m looking forward to a great new year in Sample Prep, and plan to start by going through some of the high points of the presenters made, as well as some of the highlights of our presentations there.  Personally, I think Sample Prep is the next horizon of Biotech – where we can gain some real traction and make a difference to help people around the world.  Through better, faster, and cheaper sample prep, we can all amplify the benefits accruing from the advances being made in many fields where rapid methods are being implemented.

Collection of samples for determination of genetic content can be challenging for several reasons.  The wide variety of materials present in environmental air and water means that not only organisms and “free” nucleic acids will be present, but dissolved materials, debris, interferents, and degraded genetic material will also be present.  For air, collection has to be done in a way that will allow organisms and nucleic acids to be retrieved from the sample. If the sample is collected in a liquid, it can be a little simpler than collecting on a solid substrate, such as a filter.   However, collecting into liquid presents its own challenges, because the liquid collection medium will evaporate during sampling and limit sampling time, unless a makeup supply is in place.  Collection on dry filters eliminates the requirement for makeup fluid, and generally is more convenient during sample collection.  InnovaPrep makes two “wet” sample collectors, and also makes a dry collector, the “Bobcat” air sampler.  Samples collected by the Bobcat are quickly and easily eluted into a liquid sample by using a proprietary extraction fluid and technique.  Two options for elution are in development; hand-held elutor cartridges, and a stand-alone elution station.

Once air samples for genetic analysis are captured or eluted into a liquid, it may be advantageous to further concentrate them while cleaning up the sample. This is also the case with water samples whatever the source.  InnovaPrep’s biological concentrators can be used to do that.  The HSC-40 concentrator is a bench-top instrument that allows samples of up to about a liter in volume to be concentrated in a single automated process down to 100 microliters or less for processes like gene scanning, while getting rid of ionic species and transferring the sample to a clean elution fluid.  This buffer exchange is an important step in determining genetic content of environmental samples.  Various buffers, such as Tris (tris(hydroxymethyl)aminomethane) can be used in the elution fluid and process.

The HCI-40 integratable biological concentrator is made to plug-and-play with systems or processes to concentrate particles for better detection or identification.  Input sources can include reservoirs or process streams, with the input rate limited to about 40 mL/min or less.  The output of the HCI-40 is a small preset volume of clean buffered liquid, containing the biological particles.  Output volumes of the HCI-40 can be as little as about 50 microliters.  This is accomplished using the InnovaPrep “wet-foam elution” process in combination with hollow fiber or flat fiber membrane concentration cells.  The HCI-40 is best used when repeatable, efficient, fully automated sample concentration is needed.

The HCI-40 is fairly small, about the size of a toaster.  It can fit into many existing process setups.  Typical input tubing is 1/16” Teflon or similar tubing, but it can be set up to pull from an existing sample reservoir using tubing.  The stock connector is a 24-40 thread low-holdup port to the device.  Once the operational protocol is set, the unit can be triggered remotely, by a timer, or initiated manually.  Concentration cells are good for up to hundreds of concentration cycles, and the fluid path can be decontaminated quickly using sodium hydroxide, bleach, or hydrogen peroxide.  Depending on the type of concentration cell used, particles such as viruses and DNA can be efficiently separated from bacteria and particles.  By using two HCI-40 units in cascade fashion, it is possible to separately concentrate both size components without a sample split.  That is done by processing the sample a first time, concentrating the larger particles such as bacteria, then concentrating the resulting permeate from the first unit using a second concentrator to concentrate the viruses, DNA, or other proteins.

While the SCODA technique does have its limitations, it has been shown to be effective.  A quantitative evaluation of SCODA’s ability to reject humic acids, and a comparison to existing technologies was recently demonstrated at Stanford (Cold Spring Harbor Protocols).  Distilled water samples containing 2ng of DNA were spiked with increasing concentrations of humic acids.  The samples were then purified using a commercial silica column DNA purification kit, a commercial immuno bead-based isolation system, and  Boreal’s SCODA technologies. The success of the humic acid rejection was assayed by the relative success of PCR amplification of the extracted DNA over a number of samples (ranging from 1 at the low humic acid extremes up to 20 at the relevant inflection points in the plots of the data).  It is clear from this work that SCODA was at least 100-fold better for humic acid rejection than conventional technologies.  It should also be noted that kits designed for removal of a specific contaminant will only be effective against that contaminant, while SCODA works equally well when presented with humic acids, proteins, indigo dye, hemoglobin, and other contaminants. Link: http://www.pnas.org/content/106/35/14796.figures-only  .

In summary, the InnovaPrep concentration systems can provide an efficient and rapid means for concentrating biological particles of interest into small volumes more closely matching those required when using rapid detection and identification methods.  Improvements in detection limits by orders of magnitude  can be realized.  Because the systems use size-selective based concentration, other non-target, and sometimes inhibitory, particles and large molecules may also be concentrated along with the target particles.

A wide range of inhibitor removal kits, purification kits and instruments are available or are under development by several commercial developers and suppliers.  In general, these techniques are efficient at removing inhibitors, but may require optimization for some applications.  These techniques also are nearly all more efficient and more rapid when the sample to be processed is less that 0.5 mL in volume, so they provide a good match to the InnovaPrep concentration systems that are capable of concentrating into volumes down to 50 µL.

In summary, matching the size-selective InnovaPrep technique, which is capable of accepting very large input volumes and delivering very large concentration factors, with a non-size-based separation technique that is optimized when input volumes are small, provides an elegant solution that draws on the best characteristics of both techniques.

The SCODA system is a relatively new technique, but is well proven and is currently being coupled with the InnovaPrep process in the Department of Homeland Securities’ Automated Environmental Sample Preparation (AESaP) program.  InnovaPrep and Boreal Genomics are subcontractors to ICx Biosystems under the AESP program and coupling of the two systems prior to qPCR analysis has been demonstrated.

SCODA is a nucleic acid concentration and separation method based on electrophoretic motion of molecules in a gel. It is fundamentally different from other DC and AC electrophoretic methods such as Pulsed Field Gel Electrophoresis (PFGE), in that all conventional electrophoretic separations tend to be dispersive, with bands increasing in breadth through the duration of a run. The SCODA method, on the other hand, employs a novel physical parameter associated with molecular motion in a gel to concentrate specific species of molecules – in this case, nucleic acids.  The ability of SCODA to concentrate DNA while providing efficient removal of humic substances and other inhibitors is unparalleled so far.  Downfalls of the SCODA system include the cost of the system, relatively long run times, and incompatibility with high salt concentrations.  The ability of the InnovaPrep system to concentrate samples into volumes of less than 0.5 mL and to remove salts makes coupling of these systems a powerful approach for concentration and inhibitor removal.

Some examples of demonstrated secondary clean-up methods that can be used subsequent to the InnovaPrep process include immunomagnetic separation (IMS), affinity techniques and electrophoretic techniques.  After using the InnovaPrep process to concentrate samples into volumes of 50 microliters to 1 mL, these clean-up methods can be more readily used and, as is the case with IMS, the techniques become more efficient.  In the case of electrophoretic techniques such as Boreal Genomics SCODA system, processing times can also be reduced.  Because of the extensive number of available inhibitor removal techniques, I’ll discuss only a few of most commonly used technologies and new techniques that have clear advantages when coupled with the InnovaPrep concentration systems.

Affinity-based and IMS separation techniques are the most commonly used techniques for separating target particles prior to analysis.  IMS can be used to separate whole bacteria and viruses from the InnovaPrep concentrate with resuspension into a new buffer prior to analysis.   Humic substances can reduce the efficiency of IMS, but protocols for dealing with difficult matrices (such as the IMS procedure described in EPA Method 1623) can improve the efficiency of these methods.  In general IMS is considered to be reasonably efficient in removing humics and other inhibitors and reasonably efficient in the concentrating the particles of interest into a reduced volume.  The most notable downfalls of IMS are (1) the need to use large numbers of expensive beads to produce a relatively small sample, (2) the need for relatively long incubation times, (3) relatively low recoveries and (4) the need for a separate bead type for each organism.

Affinity based methods are well proven but can suffer from overloading when large amounts of humics or other inhibitory compounds are present.  Column-based silica resin clean-up techniques, such as those offered by Qiagen, are commonly used for DNA cleanup but result in relatively low yields.  A newer affinity based technique that uses magnetic beads for separation is Invitrogen’s ChargeSwitch technology.  Positively charged magnetic beads are used to capture DNA, which binds to the bead surface at low pH.  The beads and the attached DNA are separated from the remaining sample using a magnet and wash steps are performed, in much the same way as standard IMS procedures.  Then by resuspension in a high pH buffer the beads become negatively charged and release the DNA.  This technique is proven to be efficient, but suffers from one of the downfalls of other magnetic bead based techniques when concentration is not applied first; very high concentrations of beads are required in order to allow for efficient interaction with the DNA.

Numerous other commercially available inhibitor removal techniques are available.  One of special note is the MOBIO Laboratories PowerSoil DNA Isolation Kit.  This kit was developed for preparing soil samples prior to PCR analysis.  The University of South Florida – Center for Biodefense is using this kit to clean sample concentrates prior to analysis by PCR, with good results.  In general soil preparation kits for PCR analysis are a good choice for off-the-shelf techniques because they are designed towards removal of large amounts of humics.  In addition to MOBIO, EPICENTRE Biotechnologies, Zymo Research Corporation, Omega Bio-tek Inc., and a number of other companies provide soil preparation kits.