March 2017 PhyNexus exhibits at Association of Biomolecular Resource Facilities ABRF 2017 held in San Diego, CA, USA,

PhyNexus expands its services to target academic core laboratories with automated bench top instruments. ABRF is a unique membership association comprising over 900 members with over 340 laboratories and administrative offices in government, academia, research, industry and commercial settings. PhyNexus showcases its two bench top instruments: The AutoPlasmid MEA-2 instrument for mini and midi scale plasmid purification and protein purification and the AutoPlasmid MMG instrument for maxi, mega and giga scale automated plasmid purification. The instruments are uniquely positioned to provide bench top automation while retaining the versatility of addressing virtually any protein or plasmid purification need.

February 2017 PhyNexus introduces its new automated plasmid prep MMG instrument at the Annual International SLAS Conference & Exhibition, in Washington, DC, USA

A new instrument for automated Maxi, Mega and Giga scale plasmid purifications was introduced by PhyNexus on February 4-8, 2017 at the Annual International SLAS Conference & Exhibition held in Washington, DC at the Walter E. Washington Convention Center. The Autoplasmid MMG instrument produces large scale transient transfection quality plasmids in a walk away instrument. The instrument is designed to process a wide range of cell culture densities and volumes to produce the desired amount of plasmid.

December 2016 PhyNexus exhibits at the Antibody Society’s annual meeting, Antibody Engineering & Therapeutics on December 11–15, 2016 in San Diego, CA, USA

PhyNexus displays its wide range of protein and plasmid purification automation products. The technology is based on Chromatography on a Tip chemistry for up to 96 at-at-time robotic liquid handlers to purify and process proteins, protein complexes, plasmids, and genomic DNA. Our unique back-and-forth flow of sample and buffers through the tip column produce a Tip Concentrating Effect. The process produces very high concentrations of protein even from limited sample. Samples can be grown in miniature in 96 well high throughput format saving time and money. All chemistries are available from PhyNexus and all major robotics are supported.

July 2016 PhyNexus presents Affinity Chromatography Method Development paper at PREP 2016, Philadelphia, PA, USA

The PREP conference highlights the industry shifting to a new technique called Continuous Chromatography. This technology offers higher throughput manufacturing by employing two affinity columns connected in series. As the first column is loaded, it is removed from the sample stream and replaced by the second column. A new second column is placed in the sample stream collecting any carry over from the first column. The paper presented by PhyNexus describes the ability to quickly discover the conditions that optimize rapid and complete loading and complete washing of impurities and recovery of active protein. This produces not just a pure protein but an active protein.

May 2015 PhyNexus introduces its new AutoPlasmid MEA instrument at the annual PEGS Boston held on May 2015 in the Seaport World Trade Center in Boston, MA, USA

PhyNexus introduces the new AutoPlasmid MEA for hands off bio sample prep of mini scale and midi scale plasmid purification directly from pellet. The instrument is a multi-tool bench top robotic system capable of processing 1 to 12 samples at-a-time and plates of up to 36 samples of cell pellets. The instrument also uses affinity chromatography to process and purify protein samples, especially limited volume and concentration samples. Finally, using unique gel filtration columns, desalting/buffer exchange can be performed for samples as small as 20 µL while recovering the entire sample within a few minutes.

August 2014 PhyNexus Users Group Symposium 2014 held in South San Francisco, CA, USA

PhyNexus Users Group Symposium 2014

Thank you all for your support and participation in our PhyNexus Users Group Symposium (PUGS2014) held in South San Francisco on August 27th, 2014. The symposium had great turnout with 30 different companies and institutions participating. Out of these 30, 4 of them were academic institutions, 5 were robotic instrument partnering companies, and 21 were biotech/pharmaceutical companies.

We would like to give special thanks to all 16 speakers who gave outstanding talks as well as our partnering companies (Dynamic Devices, Hamilton, Perkin Elmer, Rainin, and Tecan) for joining PhyNexus to make this symposium successful.

As requested by many of you, we now have most of the presentation slides available. Feel free to download the slides and if you have any questions, feel free to contact us or the speakers directly.

1 Douglas Gjerde PhyNexus

2 Guenther Bonn Innsbruck

3 Irina Mineyev PE

6 Bronson Duck Hamilton

7 Lee Hoang PhyNexus

8 Andras Guttman UnivDebrecen

9 Alice Yam Sutro

11 Daniel Leach Tecan

12 Rishi Porecha_Rainin

13 Tyson Bowen PacBio

14 Karen Billeci Genentech

15 Jeff Hurwitz DynamicDevices

16 Matthias Rainer Innsbruck

November 2013 PhyNexus to present poster at Advances in Recombinant Protein Technology 2013, AstraZeneca R&D, Alderley Park, UK

Advances in Recombinant Protein Technology 2013

A meeting jointly organized by AstraZeneca, University of Manchester and ELRIG

The ability to produce proteins (as reagents or biotherapeutics) of sufficient quality and quantity is important for the academic and industrial sectors. The use of eukaryotic cells as hosts for expression and recovery of recombinant cellular, membrane or secreted proteins has developed markedly over the past decade. This conference brings together academic and industrial researchers working on eukaryotic cells as protein expression hosts, and will address our current fundamental understanding about molecular events that determine productivity and quality attributes in the use of specific cell hosts for specific protein product categories.
In particular, the conference will focus on the biology of existing host cells (in terms of competencies for protein expression), structural features associated with success of expression of specific proteins (including the relationships between specific proteins and host cells) and how both host cell and desired proteins may be engineered to generate new (synthetic) systems for enhanced product expression. The two day conference will consist of presentations by plenary speakers, workshop sessions with invited speakers and speakers selected from offered abstracts and a poster session. Participation will be strictly limited to 220 attendees to enable networking and informal discussion.
Come and visit the PhyNexus poster “Completely Automated Plasmid Prep & Optimized Protein Separation Using PhyTip Columns” to learn how the PhyTip column technology provides automated, flexible and scalable solutions for low volume protein and nucleic acid purification.

Visit
http://www.elrig.org/index.php/events/152-airpt-2013/1155-advances-in-recombinant-protein-technology-2013 for details.

November 2013 PhyNexus announces the launch of the enhanced MEA 2 Personal Purification System; the next generation platform for automated chromatography applications.

SAN JOSE, Calif., November, 2013– PhyNexus Inc., announces the launch of its next generation platform, the MEA 2 Personal Purification System.
The MEA 2 is designed to process PhyTip® columns for automated and unattended purification and enrichment of soluble and membrane proteins and plasmid DNA.
The MEA 2 features a redesigned 12-channel head pipette system. This incorporates improved performance and reliability, even with viscous liquids and is suitable for cold room usage.
The back and forth active transport in the PhyTip chromatography column drives binding and elution to equilibrium maximising yield and concentration. The system is capable of processing samples sizes from 100 μL up to 80mL allowing great flexibility in a personal robotic platform. In addition to excellent purification, the ability to perform elutions in volumes as low as 10 μL provides excellent sample concentration.
Application areas that benefit from the features of the enhanced MEA 2 include process development, parallel large volume purification and crystallography sample preparation.
“We are finding that the popularity of our MEA robotic systems is increasing rapidly” says PhyNexus CEO, Douglas Gjerde, Ph.D. “Historically, our MEA line has filled an important niche for our customers. Unlike larger systems our robotic system sits on any bench top and is easy to program by our research customers who are actually using the system while still providing the automation and throughput that our customer need. We are pleased with the early success of this new MEA 2 offering.”

About PhyNexus
PhyNexus Inc. has developed a unique technology to address the need for high throughput performance for small-volume protein and plasmid purification with its range of PhyTip
columns. These columns allow for rapid and routine parallel purification in a walk-away fashion. The instruments, columns and their use is the subject of several domestic and foreign patents and patent applications. For more information, visit www.phynexus.com. PHYNEXUS and PHYTIP are registered trademarks of PhyNexus, Inc.

Contact
Tiffany Nguyen
PhyNexus, Inc., 3670 Charter Park Drive, Suite A, San Jose, CA 95136
(408) 267-7214

April & May 2013 PhyNexus to present poster at PEGS 2013 - Essential Protein & Antibody Engineering Summit, Boston, Massachusetts

PEGS 2013 – essential protein & antibody engineering summit

Cambridge Healthtech Institute is pleased to present its Ninth Annual PEGS, the essential protein engineering summit, being held this April 29-May 3, 2013 at the Seaport World Trade Center in Boston.

In addition to a new venue, the 2013 PEGS event offers six streams in the areas of protein & antibody engineering, oncology, expression, analytical, safety, and purification. These highly focused streams enable researchers to discuss and find solutions to challenges faced in the lab and hear research that has yet to be published. Conference short courses offer a primer or in-depth workshop on everything from phage and yeast display to immunogenicity and protein therapeutics. A biologics partnering forum will also be offered for both emerging companies with innovative technologies and established companies seeking collaborations.

Year after year, PEGS challenges attendees to evolve their thinking around the generation, characterization, and clinical application of proteins. Join over 1,200 of your colleagues this April in Boston to make your mark on the rapidly changing landscape of protein and antibody engineering.

Come and visit our poster “Completely Automated Plasmid Prep & Optimized Protein Separation Using PhyTip Columns” to learn how the PhyTip column technology provides automated, flexible and scaleable solutions for low volume protein and nucleic acid purification. Visit http://www.pegsummit.com/ for details.

PEGS 2013

March 2013 PhyNexus to Attend 6th Annual Proteins Congress, London, UK

PhyNexus to attend 6th Annual Proteins Congress, London, UK

The 6th Annual Proteins Congress, London, UK will attract over 200 delegates working within Proteins, Antibodies and Biotherapeutics on 25-26th March 2013.

The congress is the single most senior-level focused event in Europe, dedicated to key technological and scientific advances in the proteins industry.

With over 40 expert speakers from leading pharmaceutical, biotech, academic and research institutes the congress will address the most pressing challenges in cell line engineering, antibody drug conjugates and difficult proteins through a series of cutting-edge conference presentations.

Learn more about current developments in novel approaches to drug discovery as well as the chance to discover innovative solutions in overcoming the problematic issues surrounding immunogenicity assay development, protein aggregation and culture specific predictions.

Come and visit the PhyNeuxs booth to learn how the PhyTip technology provides automated, flexible and scaleable solutions for low volume protein and nucleic acid purification. Visit http://www.proteins-congress.com/ for details.

6th Annual Proteins Congress, London, UK

February 2013 Structural Genomics Consortium (SGC), University of Oxford presented data showing the potential of PhyTip columns to streamline protein structure determination

Structural Genomics Consortium (SGC), University of Oxford presents data showing the potential of PhyTip columns to streamline protein structure determination.

During the MipTec conference, Nicola Burgess-Brown, Biotechnology PI, gave a presentation entitled “A high-throughput baculovirus pipeline enabling structural determination of integral membrane proteins.” The talk focused on SGC’s work on integral membrane proteins and also included examples showing the potential of PhyTip columns from PhyNexus to streamline and automate its protein structure determination platform and improve its pipeline success. To view the presentation, click here.

During the PSDI (Protein Structure Determination in Industry) meeting, Georgina Berridge presented “Increasing Throughput and Characterisation of Human Proteins at the SGC.” The talk focused on SGC’s recent implementation of a high-throughput process for purifying cytosolic and integral membrane proteins (IMPs) from small scale (50 – 100 ml) E.coli and baculovirus/insect cell cultures. Purification using the PhyNexus IMAC PhyTip system has enhanced screening and enriched capture of medium to highly expressed proteins, yielding crystals in less than a week for selected targets. To view the presentation, click here.

About the Structural Genomics Consortium (SGC), University of Oxford.

The SGC is a not-for-profit, public-private partnership with the directive to carry out basic science of relevance to drug discovery. The core mandate of the SGC is to determine 3D structures on a large scale and cost-effectively; targeting human proteins of biomedical importance and proteins from human parasites that represent potential drug targets. For more information on the SGC, click here.

SGC

January 2013 PhyNexus PhyTip Normal Phase 1 Columns for High Throughput Enrichment of Glycans

PhyTip® Normal Phase 1 Columns for High Throughput Enrichment of Glycans

We have the perfect solution for high throughput, reproducible sample dye removal prior to identification and quantification of glycans.

Biopharma companies routinely characterize their antibody leads and glycoproteins through analyses such as glycan mapping. PhyTip Normal Phase 1 Resin columns are designed for the specific enrichment of polar compounds. These PhyTip columns are especially useful in glycan mapping and identification analysis of glycoproteins. High resolution tools such as HPLC and capillary electrophoresis are required for the separations of these complicated structures; however, detection must be enhanced through use of fluorescent dye labels. PhyTip Normal Phase 1 Resin columns are the perfect solution for high throughput, reproducible sample dye removal prior to identification and quantification.

PhyTip Normal Phase 1 Resin column features include:

  •  Separation and enrichment of polar compounds such as glycans.
  • Excellent solution for excess dye removal prior to CE or HPLC analysis of glycans.
  • High throughput, automatable glycan purification solution.
  • Processing of 12 samples in parallel with the MEA Purification System or 96 samples in parallel using standard liquid handling robots.
  • Standard column sizes:
    • 200 µL column containing 5 or 20 µL of resin.
    • 1000 µL column containing 10 or 20 µL of resin.

Reasons why others use PhyTip Normal Phase 1 Resin columns:

  • ‘Use of paper chromatography, whilst effective, is hindered by the time commitment and reduced efficiency in dye removal that is readily evident in subsequent NP-HPLC separation. So we looked for an alternative that would be quicker, more efficient and preferably automatable. PhyTip normal phase 1 columns work extremely well. The benefits are speed, efficiency of excess dye removal and ease of use.’ Dr Ciara McManus, NIBRT
  • Publications citing PhyTip Normal Phase 1 Resin columns include:
    • Bradley D. Prater *,1, Kalyan R. Anumula, Jeff T. Hutchins, Automated sample preparation facilitated by PhyNexus MEA purification system for oligosaccharide mapping of glycoproteins. Analytical Biochemistry 369 (2007) 202–209.
    • Bradley D. Prater *, Heather M. Connelly, Qiang Qin, Steven L. Cockrill, Highthroughput immunoglobulin G N-glycan characterization using rapid resolution reverse-phase chromatography tandem mass spectrometry. Analytical Biochemistry 385 (2009) 69–79
    • Zoltan Szabo, András Guttman, and Barry L. Karger*, Rapid Release of N-Linked glycans from glycoproteins by pressure cycling technology. Anal Chem. 2010 March 15; 82(6): 2588–2593.

For more information:

September 2012 PhyNexus Introduces New Wider-Based Guidelines for Culture Preparation of Plasmids for Plasmid Purification

Plasmid Purification News

New wider-based guidelines for plasmid purification have been released by PhyNexus. The guidelines describe how to prepare samples to streamline the Lysate Direct PhyTip column method for Plasmid DNA purification starting from E. coli cell pellets. The new guidelines demonstrate completely automated, Direct-From-Pellet plasmid technology that can be used for medium and high copy plasmid construct to yield high-quality plasmid. Small volume cultures can be grown directly in a 96 well format with high efficiency for everyday laboratory needs such as transfection, sequencing and PCR.

The guidelines start with describing recommended procedures for growing the culture. The plasmid culture cells may be several different strains including DH5α, DH10, TOP10, Mach 1, XL1-Blue, XL2-Blue or similar type cells that are suitable for transformation of plasmid and ligated material. Ultracompetent cells are also recommended for transformation of large clones.

Plasmids fall into three general categories: High, Medium and Low copy corresponding to the copy number per cell. Cloning vectors used may vary depending upon the cloning method and features of the plasmid including origin of replication and antibiotic resistance. Expression vectors will exhibit lower copy number and will be specific for the expression system. It is recommended that low copy plasmids be cloned to medium and high copy vectors to obtain higher yields when possible.

The guidelines also discuss the antibiotic resistance markers, inoculations procedures, culture plate seals, shaker, centrifuge conditions and media. The media in particular can be important to establish high growth conditions and plasmid yield.

Call PhyNexus at 408.267.7214 or email [email protected] to request your copy of the Guidelines.

Plasmid Purification News

September 2012 Rainin Mettler Toledo Introduces Fast Immunoprecipitation in a Pipette Tip

Fast Immunoprecipitation in a Pipette Tip

Quickly isolate specific proteins from cellular lysate, serum or other biological fluids with Rainin’s new PureSpeed Protein Purification System. PureSpeed is a unique approach to protein purification offering simultaneous processing of up to 12 samples. Rainin’s new PureSpeed Protein Tips, in concert with its pre-programmed E4 XLS electronic pipettes, deliver highly concentrated immunoprecipitated proteins with very low background in less than 30 minutes.

More information can be found at http://www.phynexus.com/automation-workflow-integration/

PipetteTip

July 2009 Caliper Life Sciences and PhyNexus Held Protein & Genomics Conference

PhyNexus in collaboration with Caliper Life Sciences held a Protein & Genomics Dual Conference in South San Francisco and in La Jolla, California

July, 2009 – PhyNexus, Inc. in collaboration with Caliper Life Sciences announced the Protein & Genomics Dual Conference held at the Embassy Suites Hotel in South San Francisco on June 2nd and Hilton La Jolla Torrey Pines on June 4th. Presentations on high throughput solutions for protein and nucleic acid purification, protein production, protein engineering, cell-based assays, and bioprocess development were delivered by researchers from various Biopharmaceutical companies. The conferences were well attended by both researchers from Biopharmaceutical industry and academia. Over 200 researchers attended the conferences and participated in talks, presentations, new product demo as well as social events.

Among many interesting talks, Dr. Lee Hoang from PhyNexus delivered a talk titled “Automated, high throughput protein purification and sample prep using PhyTip columns for therapeutics leads screening and process development.” In his talk, Dr. Hoang highlighted the advantages of using PhyTip columns which can relieve bottlenecks in therapeutic development process by increasing throughput, incorporating automation, and providing reliable and robust purification solutions.

PhyTip columns are compatible with the Caliper Sciclone and Zephyr liquid handling instruments for the parallel processing of 96 samples. Generation of these many samples requires a reliable and efficient means of analysis. The Caliper LabChip GXII offers automated analysis, and quantitation of molecular weight, concentration and purity.

January 2008 Screening for Chromatographic Success and Failure Presentation at Cambridge Healthtech Institute Peptide and Protein-Based Therapeutics.

Screening for Chromatographic Success and Failure on Small Scale

William Gillette, Ph.D., Senior Scientist, Protein Purification Group Leader, Protein Expression Laboratory, Advanced Technology Program, NCI/SAIC-Frederick

For proteomic and core service laboratories charged with purifying numerous proteins, a method to quickly screen the outcomes of different chromatographic approaches for multiple proteins in small-scale would be extremely useful in terms of reducing costs, increasing throughput and, more importantly, improving the success rate. Such a method must have the flexibility to interrogate multiple parameters, while also providing enough material for downstream analysis. We have approached this problem by maximizing the usefulness of samples which are generated by the expression screening groups of our lab and minimizing the amount of sample required for the purification screen.

4th Annual Cambridge Healthtech Institutes

Peptide and Protein-Based Therapeutics

January 7-9, 2008 in San Diego, California

January 2008 Using DOE and Microscale Techniques to Define Process Boundaries Presentation Well Characterized Biotechnology Pharmaceutical Symposium.

Space is the Place: Using DOE and Microscale Techniques to Define Process Boundaries

Marc Wenger

Merck & Co., Inc., West Point, PA USA

Development of protein purification processes is a multivariate challenge to select a winning combination of unit operations and operating parameters that maximize purity, recovery, and process robustness while minimizing cost. For the development of de novo processes, this is a time-consuming and assay-intensive exercise. Consequently, platform purification processes have emerged as a means of streamlining process development for proteins having similar physicochemical properties. However, despite having fixed unit operations, platform purifications still require some optimization to adjust for differences in molecular properties between proteins. Techniques for miniaturizing protein purification processes at the microliter scale (microscale) can accelerate process development and reduce resources by enabling parallel experimentation and automation. Design of experiment (DOE) methodologies provide an additional tool for examining multiple variables simultaneously, and when combined with microscale techniques, enable purification processes to be rapidly and efficiently optimized. Two examples will be shown in which DOE in combination with microscale methods was used to optimize process conditions and define operating boundaries. In the first example, factorial and response surface designs were used to screen and optimize the parameters for a small-scale disruption of yeast cells. In the second example, the operating space for two ion-exchange chromatographic steps in a platform purification were defined for a monoclonal antibody, resulting in improved process performance and robustness.

Well Characterized Biotechnology Pharmaceutical Symposium

California Separation Science Society

January 27-30, 2008 Washington DC

April 2007 PhyNexus wins Best Poster Presentation at 13th Annual SBS Conference and Exhibition in Montreal, Quebec, Canada

January 2007 Optimization of Protein Separation Using Automated Micro-Scale Column Technology.

If you are attending the SBS Conference in Montreal and would like to discuss the PhyNexus products or technology in more detail, please contact us at [email protected] and we will make arrangements for you to meet with a representative from PhyNexus.

Society for Biomolecular Screening
8th Annual Conference and Exhibition

April 15-19, 2007
Montreal, Canada
Poster No. P10003

Session: Structural Biology: Underpinning Drug Discovery Title
“Optimization of Protein Separation Using Automated Micro-Scale Column Technology”

January 2006 Cambridge Antibody Technology gives presentation at Lab Automation Conference in Palm Springs.

January 2006 PhyNexus gives presentation at CHI’s PEP Talk in Coronado, CA.

August 2005 PhyNexus Participation in the "Developments in Protein Interaction Analysis" Symposium sponsored by Biacore in Philadelphia, PA.

August 2005 PhyNexus and Raven give poster presentation at IBC’s Drug Discovery Conference in Boston, MA.

IBC’s Drug Discovery Technology Poster, August 8-11, 2005, Boston MA, Boston Convention and Exhibition Center

Automated enhancement and streamlining of therapeutic antibody discovery through the application of micro-scale high-performance protein separation technology

Chris Hanna, Doug Gjerde and Jeremy Lambert, PhyNexus, Inc. Tony Liang and John Whelan, Raven Biotechnologies, Inc.

Due to the intrinsic quality that monoclonal antibodies have for highly targeted interactions with their antigens, their success as anti-cancer therapeutics has been particularly notable. A significant challenge associated with the discovery of these antibodies is the identification of drug targets that yield to antibody intervention, along with identification of corresponding antibodies that confer the desired functional outcome upon interaction with the target. While many technologies have evolved over the past decade to achieve these tasks, one particularly notable approach allows for simultaneous identification of both drugable cell-surface targets that possess true functional specificity and identification of monoclonal antibodies that modulate target function in such ways that make them attractive drug leads. By applying novel technologies for fully automated micro-scale high-performance purification and enrichment of the candidate antibodies, along with co-purification and enrichment of the antibodies’ interaction partners, certain key tasks within this drug discovery platform can be enhanced and streamlined. This presentation will describe the application of these separation technologies to this simultaneous target/antibody discovery platform, the specific enhancements provided by doing so, and the overall improvements brought by this separation technology to therapeutic antibody discovery.

June 2005 PhyNexus gives Poster Presentation at CHI’s "Beyond Genome" Proteomics Conference in San Francisco, CA

CHI’s “Beyond Genome” Proteomics Conference Poster, June 15-16, 2005 San Francisco, CA, Fairmont Hotel

Automation of High-Performance Micro-Scale Protein Interaction Analyses

Christopher Hanna, Jeremy Lambert and Douglas Gjerde, PhyNexus, Inc, San Jose, CA Jeffrey Sklar and Hui Li, Yale University School of Medicine, New Haven, CT Tony Liang, Claudia Fieger and Irene Ni, Raven Biotechnologies, South San Francisco, CA David A. Herold and Lynn Nisbet, University of California, San Diego, CA

In the post-genome era increasingly greater attention is given to deeper elucidation of gene function. This is particularly the case for the discovery and development of successful disease diagnostics and therapeutics. For example, ever-increasing proteomics-driven searches for diagnostic biomarkers and potential therapeutic targets demonstrate the need for identifying integral functional players within the disease itself. Once a putative biomarker or drug target within a disease process is identified it’s proper place and role within the larger disease pathway must be determined, as well as determining the factors that regulate the presence and absence of this presumed functional player within the disease state. Various techniques and sub-techniques for the analysis of protein interactions are typically applied for these determinations, and many suffer from poor performance characteristics – e.g., they are difficult to automate and often labor-intensive, they possess poor enrichment and purification characteristics, they require overly large samples, etc.

A novel micro-scale affinity column technology is described that enables many of the most critical techniques for protein interaction analyses – such as immunoprecipitation (IP), GST pulldowns, coimmunoprecipitation (CoIP) and chromatin immunoprecipitation (ChIP) – to be performed in an easily automated and straightforward manner. The amount of sample required for successful analyses is a fraction of what is typically required, and performance characteristics such as sensitivity and purity are often dramatically improved over standard techniques. The technology can also be integrated directly into existing fluidic automation, thus opening the door to fully automated high-performance protein interaction analyses for 96 samples at a time.

June 2005 Presentation at Academy of Clinical Laboratory Physicians and Scientists 40th Annual Meeting, Pittsburgh, PA

ACADEMY OF CLINICAL LABORATORY PHYSICIANS AND SCIENTISTS 2005

(2005-##) Transferrin Extraction By Immunoaffinity Microcolumn Chromatography. Joan E. Bernard1 , Lynn Nisbet1 , Fe Sumabat1 , David A. Herold*1 , Jerald S. Feitelson2 , Lei Huang2 , Christopher Hanna3 , Doug Gjerde3 . 1 Departments of Pathology, University of California, San Diego, La Jolla, CA 92037 and Veteran Affairs Medical Center San Diego, CA 92161, 2 GenWay Biotech, Inc., San Diego, CA 92121, 3 PhyNexus, San Jose, CA 95136.

Background: Transferrin has traditionally aided clinicians in the diagnosis of iron-deficiency anemia. More recently, carbohydrate-deficient transferrin isoforms have been used for monitoring ethanol abuse. In this study, we used 200 microliter PhyTip™ immunoaffinity microcolumns (IAMC) containing 5 microliters of immobilized IgY (avian) antibody coated microbeads (Seppro™) to selectively remove transferrin from serum for further analysis.

Objective: Establish proof of principle by recovery of transferrin from serum using IAMC.

Methods: Serum transferrins were measured by the standard Beckman LX20 turbidimetric assay in 44 patients. Using these samples, serum transferrin was isolated by IAMC and concentrations were measured using a Beckman Access turbidimetric assay adapted for the LX20. Results for the two methods were used to determine the recovery of transferrin from serum using the IAMC. In addition, the potential for regeneration and reuse of the IAMC was evaluated. SDS-PAGE was used to verify transferrin isolation.

Results: Transferrin concentrations in three patient samples by the LX20 turbidimetric assay were 127, 229 and 331 mg/dL (n=6). Their recoveries measured by the modified Access assay after IAMC separation using new columns (n=6) were 62 +/- 5%, 80 +/- 4% and 78 +/- 6%, respectively. Additionally, recoveries when reusing the same columns 6 times were 79 +/- 16%, 86 +/- 17% and 85 +/-15%, respectively. The isolation of >90% pure transferrin was confirmed using SDS-PAGE. Subsequently, transferrin was isolated in 44 patient samples using the IAMC technique. The comparison of transferrin by column capture versus standard serum turbidimetric assay yielded an average recovery of 91% and an overall R2 = 0.78 for serum values ranging from 60 to 330 mg/dL.

Conclusion: IAMCs can be used to specifically isolate transferrin from other serum proteins and could be applied to the measurement of other serum proteins using appropriate IgY antibody-coupled microbeads. Further investigation of this technique may have applications in such areas as the measurement of carbohydrate-deficient transferrin, a novel biomarker for ethanol abuse. While there was some randomness in the recovery obtained, there was no appreciable decline in transferrin recoveries after six uses for each of five different columns.

May 2005 PhyNexus gives Poster Presentation at CHI’s Phage Display and Recombinant Antibodies Technology

CHI’s Phage Display and Recombinant Antibodies Technology Poster, May 16-20, 2005, Cambridge, MA, Royal Sonesta Hotel

Novel strategies for assaying recombinant antibody function with high-throughput cell-based assays

C. Hanna, D. Gjerde, D. Burge, L. Nguyen and J. Lambert, PhyNexus, Inc. San Jose, CA T. Arvedson, Amgen, Inc. Thousand Oaks, CA

Phage and related display technologies enable screening of vast repertoires of recombinant human antibodies for isolation of cognate binders to putative disease targets. Screening operations are typically performed on the basis of antibody-target affinity, with subsequent winnowing of candidates on the basis of other antibody-target binding characteristics (such as its ability to prevent native ligand binding). Once the number of antibody candidates is substantially reduced, more descriptive biofunctional data – which ultimately provides the biological basis of antibody potency – are obtained from more descriptive cell-based assays.

As it has been demonstrated that antibody features such as affinity provide a highly unreliable proxy for antibody potency1-3, there is growing need to gain more descriptive potency-correlated (i.e. cell-based) data on individual antibody clones earlier in the antibody discovery process. However, there are a number of factors that confound attempts to obtain this data, particularly in the earliest stages of E.colibased antibody discovery. For example, the direct analysis of E.coli-derived recombinant antibody subclones is confounded by endotoxins and other contaminants intrinsic to the gram-negative bacteria used in the antibody expression process. In addition, the concentrations of antibody present in E.coli preparations are typically of insufficient concentration for use in most cell-based assays. These limitations are most frequently addressed by scaling up the individual growths and using larger-format protein separation and purification tools that do not lend themselves to high-throughput automation – thus creating the present situation where cell-based assays are performed later in the discovery process.

PhyNexus, Inc. PhyTip™ column technology is applied as a high-throughput means for purification and enrichment of recombinant antibodies from small-scale E.coli sub-clone preparations that eliminate the need for scale-up, and thus enable true high-throughput cell-based assays of recombinant antibody function. This capability is demonstrated for cell-based assays of antibody antagonist activity via Statdependent luciferase reporting of Stat5 phosphorylation. Factors that contribute to PhyTip column and cell-based assay performance are discussed and include column flow and elution conditions, chemical composition of eluents, and optimizing these conditions for the maintenance of cellular viability.

1. G.P. Adams, R. Schier, A.M. McCall, H.H. Simmons, E.M. Horak, R.K. Alpaugh, J.D. Marks and L.M.Weiner, Cancer Res., 2001, 61, 4750-4755.

2. C.P. Graff and K.D. Wittrup, Cancer Res., 2003, 63, 1288-1296.

3. D. Pohlers, C.B. Schmidt-Weber, A. Franch, J. Kuhlmann, R. Brauer, F. Emmrich, R.W. Kinne, Arthritis Res., 2002, 4, 184-189.

April 2005 PhyTip columns feature in April 7 issue of Nature Methods

The recent Technology Feature in Nature (Vol. 434, pp 795-798, April 7, 2005) included a section titled “Small-Scale Separation” that featured PhyTip columns and how PhyNexus is meeting the demand for micro volume high performance protein purification.

With PhyTip columns, there is a high trapping efficiency for the sample. A 10-20-fold increase in the target protein sample can be obtained from just a few hundred microliters of sample, and purities are often over 95% with a single separation step. PhyTip columns are compatible with the automated PhyNexus ME and MEA systems plus a number of commercially available liquid-handling robots.

April 2005 PhyTip plate covers to reproducibly purify and enrich recombinant proteins and antibodies

By using PhyTip plate covers, evaporation rates are significantly reduced allowing for unattended operation of the purification process and producing reproducible, quantitative data over the complete multiwell plate.

TechNote PHYTIP ™ COVERS IN CAPTURE PURIFY ENRICH PROCESS

March 2005 New MEA Purification System for fully automated purification of up to 96 samples

The MEA Purification System offers a range of flow rates that maximizes the purification and enrichment efficiency of the various affinity resins available for PhyTip columns. This unique system allows for reproducible positioning of the PhyTip columns into the sample solution, wash buffer(s) and finally elution buffer used for enrichment. By maintaining the optimum position, the system is able to accurately deliver the programmed volumes of liquids through each column.

PHYTIP™ MEA PURIFICATION SYSTEM FOR AUTOMATED PHYTIP™ COLUMN SAMPLE PROCESSING

The Personal Purification System, comprising of the MEA Purification System and PhyTip columns, allows researchers to rapidly perform high performance micro scale purifications of their recombinant proteins and antibodies. The System is ideal for purification of recombinant proteins and antibodies ready for functional analysis, optimizing _expression conditions within process development, and optimizing purification conditions prior to scale-up.

PERSONAL PURIFICATION SYSTEM

January 2005 PhyTip columns feature in Nature Methods

The recent Technology Feature in Nature Methods (Vol. 2 No. 1, January 2005) included a section titled “Striving for purity: advances in protein purification” that featured PhyTip columns and how PhyNexus is meeting the growing demand for micro volume protein purification.

PhyNexus’ novel approach to purification incorporates affinity resins into the 200 µL and 1,000 µL PhyTip columns. Users have a choice of manual, semi automated, fully automated or full robotics platforms to perform separations. The PhyTip columns are capable of high purification (>95%) and enrichment (up to 5 mg/ml) of fully functional proteins.

January 2005 PhyTip Presentations at CHI’s PEP Talk

Dr. Chris Hanna, Executive VP of Business Development presented twice at the CHI PEP Talk – The Protein Information Week, San Diego, January 10 to 13, 2005.

On January 10th, Hanna was a co-presenter at the Buzz Talk session on High Throughput Expression and Purification along with Mark Roskey of Caliper.

Are Your High Throughput Expression Options Limited by Downstream Processing?

On January 12th, Hanna presented a paper entitled, “Unique enabling tools for protein process optimization, are your high throughput expression options limited by downstream processing?”

Unique enabling tools for protein process optimization

January 2005 PhyNexus and Caliper Life Sciences give joint Poster Presentation at CHI’s PEP Talk

PhyNexus presented a joint poster with Caliper Life Sciences at the recent PEP talk titled, “Improved process efficiency with 96-well protein purification and characterization.” In this poster, PhyNexus presented an integrated approach that included the automated purification of micro volume protein samples in a 96-well format along with real time monitoring of the resulting purity and concentration data.

The research focused on the simultaneous purification and enrichment of up to 96 samples in under 15 minutes. This was performed by automated selective enrichment of monoclonal antibodies from hybridoma growth media and recombinant 6X-His or GST-tagged fusion proteins from E. coli lysate using PhyTip columns on the Sciclone ALH 3000 workstation. Subsequent characterization (size, concentration, purity) of the enriched material using the LabChip 90 microfluidic protein analysis system provides a streamlined workflow and significant reduction in time and labor associated with high throughput protein expression and purification. The purity and concentration of the final products enable downstream analytical and biochemical assays, increasing productivity and decreasing development time for therapeutic antibody drugs.

January 2005 Joint publication between PhyNexus and Dyax appears in Jan 1 issue of Genetic Engineering News

Vol 25 No 1 of Genetic Engineering News, published on January 1, 2005, contained a tutorial paper co-authored with scientists from Dyax Corporation titled, “Antibody Enrichment and Characterization – utilizing the PhyTip Sample Preparation System.”

PhyNexus has developed a novel sample preparation technology that can rapidly purify and enrich IgG and Fab samples at a micro volume scale, where the final sample is ready for sensitive kinetic analysis using high throughput surface plasmon resonance. PhyNexus is able to achieve high throughput antibody affinity screening through the combination of improved enrichment capability and compatibility with multiple fluid handling formats. With this enabling technology researchers will be able to accelerate the identification of high quality human antibody leads in drug discovery.

December 2004 PhyNexus presents joint poster with Caliper Life Sciences at IBC’s 15th annual international conference on Antibody Engineering San Diego, CA

PhyNexus presented a poster titled: Improved process efficiency with 96-well protein purification and characterization, a joint presentation with Caliper Life Sciences that detailed the use of the Caliper LH 3000, PhyTip columns and Caliper LC 90 detection system for high throughput purification, enrichment and detection of Antibodies and recombinant proteins.

During the scientific session PhyTip products were featured twice in talks by Dr Dev Sidhu of Genentech and Dr Clive wood of Dyax:

Dr Dev Sidhu of Genentech discussed his research involving phage-displayed antibody libraries with synthetic complementary determining regions. The purpose of the work was to investigate the basic principals of antigen recognition using libraries containing limited diversity. These libraries could lead to the generation of new antibody reagents and therapeutics. Dr Sidhu used micro scale PhyTip columns for high throughput purification and enrichment of Fabs enabling him to produce high concentrations of the functional proteins for assay by SPR. This work required the generation of large numbers of samples and the problem of expressing and then purfying these samples was solved by the use of PhyTip technology. PhyTips enabled the high throughput process of rapidly purifying 96 samples at a time, producing high concentrations of functional protein that could be assayed directly, speeding up the whole discovery process.

Dr Clive Wood discussed the process developed at Dyax for Antibody drug discovery which requires a rapid and reliable means to identify high-affinity leads. In combination with automated selection and screening methodologies, thousands of Fab hits can be examined and in multiple projects, this has led to the rapid identification of very potent and selective Fab leads without in vitro affinity maturation. Dr Wood reviewed the use of PhyTip columns in their process of high throughput purification. PhyTips columns were necessary for the parallel processing of 96 samples at a time producing high concentrations of functional Fabs that could be assayed directly with the high throughput SPR system from HTS Biosystems.

December 2004 Caliper Life Sciences and PhyNexus Announce Complete Solution for Rapid, Automated Antibody Purification and Analysis

San Jose, California

December 1, 2004

Caliper Life Sciences and PhyNexus Announce Complete Solution for Rapid, Automated Antibody Purification and Analysis

HOPKINTON, Mass. and SAN JOSE, Calif., November, 2004 — Caliper Life Sciences, Inc. (Nasdaq: CALP), in collaboration with PhyNexus Inc, announced that it has integrated the PhyNexus® PhyTip™ column technology for protein purification and enrichment into the Caliper Sciclone ALH 3000 Liquid Handling Workstation, followed by rapid detection with the LabChip 90® electrophoresis system. In as little as 15 minutes, researchers can purify and enrich up to 96 engineered proteins or antibodies, and confirm purity and yield in about a minute per sample.

“We are excited to offer our customers new turn-key solutions that combine our leading expertise in liquid handling and LabChip technologies with the proprietary PhyNexus products that complement our focus on high throughput sample preparation for genomic and proteomic research,” commented Mark Roskey, Ph.D., Vice President of Marketing at Caliper Life Sciences. “The ability to couple automated high-throughput protein purification with rapid, high quality separation and analysis on the LabChip 90 system will truly enhance standardization and efficiencies of any protein research group or production facility.”

“By integrating the combined technologies available from Caliper and the novel high performance purification products from PhyNexus, our customers who are involved with antibody therapeutics will be able to cost-effectively increase their throughput capabilities and hence reduce the time to screen for potential antibodies that can be brought to trials,” said PhyNexus Executive Vice President of Business Development, Christopher Hanna, Ph.D.

About the Caliper Sciclone ALH 3000 Liquid Handling Workstation and LabChip 90 Electrophoresis System

The Caliper Sciclone ALH 3000 Liquid Handling Workstation offers leading-edge precision, adaptability, and scalability, and enables the automated processing of reagent samples for life science research and diagnostic testing. Leveraging a unique, open and modular design along with a wide range of interchangeable pipetting options and accessories, the Caliper Sciclone Liquid Handling Workstation can be configured to automate many applications, from nucleic acid and protein sample preparation protocols to molecular and cell-based screening assays, and more.

The LabChip 90 System offers complete protein and DNA analysis without the trouble of SDS-PAGE and agarose gels. The system samples directly from 96- and 384-well plates and automates the manual steps of slab gel electrophoresis onto a microfluidic chip. Quantitative sizing and concentration data are generated in just seconds per sample, and hundreds of samples can be processed in a few hours. This inexpensive alternative to gels is ideal for analysis of crude lysates, purified proteins, PCR products, restriction digests and more.

About PhyTip columns

PhyTip columns (Patents Pending) are unique proprietary devices designed to capture, purify and enrich antibodies and engineered proteins using affinity resins such as Protein A, Protein G, IMAC (Ni-NTA) and Glutathione. The process to purify(>95%purity) and enrich samples (up to 50x enrichment) in a micro volume format, can take less than 15 minutes and produces a fully functional protein for use in subsequent functional assays.

About Caliper Life Sciences

Caliper Life Sciences uses its advanced liquid handling and LabChip technologies to create leading-edge tools that accelerate drug discovery and enable diagnosis of disease. Caliper headquarters is located in Hopkinton, Massachusetts, with R&D, operations and manufacturing facilities for LabChip devices in Mountain View, California, and direct sales, service and applications support throughout the world. Caliper customers and partners include many of the largest pharmaceutical, biotechnology, and life sciences companies. For more information, please visit Caliper’s web site at www.caliperLS.com.

About PhyNexus

PhyNexus has developed a unique technology to address the need for high performance, micro volume protein purification and enrichment with its PhyTip columns. These disposable devices can be used to rapidly prepare fully functional, purified recombinant proteins and antibodies with the highest performance. PhyNexus is headquartered in San Jose, California. Further information on PhyNexus can be found at www.phynexus.com.

LabChip and Caliper are registered trademarks of Caliper Life Sciences, Inc.

PhyNexus is a registered trademark of PhyNexus, Inc.

PhyTip is a trademark of PhyNexus, Inc.

November 2004 PhyNexus launches new enhanced website features for easy ordering

PhyNexus has introduced a new feature to its current website that allows users to not only learn about the current products, but also to order them directly from the internet. With this new capability, users will be able to simply scan for the product that fits their need and order immediately. The PhyNexus Shopping Cart page employs the SmartCart™ Ecommerce software, in which users can easily navigate to search for products and order by credit card at any time.

October 2004 PhyNexus company profile in October issue of American Biotechnology Lab

PhyNexus is a company dedicated to advancing the practice of sample preparation in the life science laboratory. By developing expertise in the areas of small-scale liquid handling and chemical affinity, it has launched products that efficiently purify and concentrate protein samples with volumes of less than 500 µL and full retention of function. PhyNexus is committed to the development of an expanding range of purification and enrichment products based on traditional and novel chemistries. The company is headed up by CEO Douglas T. Gjerde and Executive Vice President Christopher P. Hanna.

Company Profile

September 2004 PhyNexus presents joint poster with Caliper Life Sciences at the Society for Biomolecular Screening’s 10th Annual Conference in Orlando, Florida

PhyNexus and Caliper Life Sciences presented a joint poster titled, “A novel approach to automated high throughput protein enrichment and characterization.”

The poster described a unique, yet simple process to purify and enrich antibodies using a column in a tip approach. These PhyTip columns were used in a fully automated 96-well system, and are capable of purifying protein from complex samples (cell culture supernatant). The purities (>95%) and concentrations (>5mg/mL possible) of the enriched proteins were characterized by real-time microfluidic technology.

September 2004 PhyNexus, Inc and QIAGEN N.V licensing agreement

San Jose, California, September 28, 2004

PhyNexus, Inc., a leading supplier of micro volume protein purification systems and QIAGEN N.V. (NASDAQ: QGENF; Frankfurt, Prime Standard: QIA) a leading supplier of products and technologies for biomolecule separation, purification and handling today announced the signing of a licensing agreement.

QIAGEN, as the exclusive license holder of patents for the His-tag technology granted to F. Hoffmann-La Roche, is manufacturing and selling 6xHis-tag coding vectors and Ni-NTA resin into the research market. Under these patent rights QIAGEN has granted PhyNexus the right to incorporate QIAGEN’s Ni-NTA resin for the purification of His-tagged protein into its patent pending PhyTip™ columns which are specifically designed for micro volume protein purification and enrichment.

“The addition of Ni-NTA as an affinity resin to capture His-tagged proteins will allow PhyNexus to expand its existing portfolio of products for micro volume protein purification and provide researchers with a unique tool for high performance purification”, said Douglas Gjerde, CEO of PhyNexus. “PhyTip columns are capable of very high purification (>95%) and enrichment (up to 5 mg/mL) using a variety of different affinity resins in a range of formats from one to 96 at a time, and maintain the functionality of the purified protein. With Ni-NTA based PhyTip columns, researchers will now be able to purify His-tagged proteins at a micro volume scale which has never been possible before.”

About PhyNexus.

PhyNexus has developed a unique technology to address the need for high performance, micro volume protein purification and enrichment with its PhyTip columns. These disposable devices can be used to rapidly prepare fully functional, purified recombinant proteins and antibodies with high enrichment. PhyNexus is headquartered in San Jose, California. Further information on PhyNexus can be found at www.phynexus.com.

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