Laboratory Focus July 2012 by Promotive Communications

Pharmaceutical

Clinical

Chemical

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w w w. b i o s c i e n c e w o r l d . c a

New Bioanalytical Tools and Devices Chemistry leads the way Page 7

e n v i r o n m e nt

July 2012 Volume 16, Number 4

Micro and nanostructured materials Page 13

R&D News.......................... 1 Appointments..................... 5 Pharma Notes..................... 6 New Products................... 15 Calendar........................... 17 Career Spotlight............... 18

Janelle Tam, 1st place award winner Sanofi BioGENEius Challenge Canada competition

At the 2012 International BioGENEius Challenge, held in Boston, Canada was well represented with projects from teen scientists Janelle Tam, a 16-year-old from Waterloo Collegiate Institute, and Rui Song, a 16-year-old from Walter Murray Collegiate, in Saskatoon. Teen scientist Song went on to place third at the international

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competition for her research on more nourishing lentils, an important global food source and one of Saskatchewan’s main crops. The competition was for high school students who through science research projects show above-average understanding of biotechnology. Winners of the competition were announced dur-

Rui Song 2nd place in Sanofi BioGENEius Challenge Canada competition

ing the keynote luncheon at the 2012 Bio International Convention. Song’s mentors, Dr. Kirstin Bett and Rob Stonehouse of the Plant Sciences Department at the University of Saskatchewan said her work raises the hope of developing a new, more nutritious variety of lentil. “As a participant since 2008, my SBCC experience has definitely changed my life,” said Song in a release. “Not only did I receive a glimpse in the research process, I gained a new perspective on career opportunities in the biotechnology sector. My experience has shaped my future career path and motivated me to change the world for the better through research.” Canada’s other competitor,

Janelle Tam, who took first place at the national 2012 Sanofi BioGENEius Challenge Canada in May, was awarded a $500 prize and honourable mention at the international competition for her project demonstrating that cellulose is also a potent anti-aging disease-fighting compound. Her mentor was Dr. Zhaoling Yao from the University of Waterloo. Tam’s research could potentially improve health and antiaging products, including tablets, bandages or cosmetic cream. “In research, I get to discover what no one has found out before, which is really exciting,” said Tam, in a press release. “I think this opens up a whole new field for NCC, and I’m thrilled that the Continued on page 3

Photos courtesy of the National Research Council of Canada

Two Canadian teen scientists compete at BIO2012 international biotech competition

July 2012 Laboratory Focus www.bioscienceworld.ca

news Encycle Therapeutics launches ground-breaking chemistry platform

Continued on page 4

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to medicinal chemists. Encycle’s chemistry platform stabilizes linear peptides for drug development by cyclizing them. This process has demonstrably increased the drug-like properties of these

tn thean ppe n cieonce t ce scDedic

ced p an ou nee ro y h g in dt yth o mdu Evn r e

metabolic disorders. In its linear form, however, peptides are unstable. When used as drug-delivery agents, they break down before reaching therapeutic targets or fail to permeate cell walls, presenting a long-standing challenge

d s cien

Peptides have long been sought after as therapeutics due to their high specificity — they can hit specific cellular targets, especially complex protein-protein interaction targets implicated in cancer, cardiovascular disease and

Encycle Therapeutics, a MaRS Innovation spin-off company, has created a ground-breaking chemistry platform for cyclizing peptides that promises to increase the efficiency of the early-stage drug discovery process.

3/20/2012 2:36:00 PM

www.bioscienceworld.ca Laboratory Focus

July 2012

news

Continued from page 1

Toronto’s Hospital for Sick Children adopts Life Technologies’ Ion Proton™ sequencer to launch new Centre for Genetic Medicine

The Hospital for Sick Children (SickKids) is partnering with California based Life Technologies Corporation to advance pediatric genomic research. Under the agreement, the semiconductor-based platform Ion Proton™ Sequencer will be the primary instrument on which multiple clinical research samples will be mapped daily in the hospital’s newly launched Centre for Genetic Medicine. SickKids and Life Technologies will collaborate on developing sequencing workflows and protocols for the Ion Proton™ System that will be tailored for studies of interest to researchers in the Centre. The first collaborative project will focus on sequencing clinical research samples to better understand the genetics behind autism, with a long-term goal to sequence up to 10,000 genomes per year to study various diseases in children. “The perfect storm of unparalleled advances in genome sequencing technology and information science, and a captivated hospital striving for new ways to move forward in medical treatment, bring us to this important day,” says the new Centre’s co-director, Dr. Stephen Scherer, who also leads The Centre for Applied Genomics at SickKids and the Univer-

sity of Toronto’s McLaughlin Centre. “We are very excited to work with Life Technologies to enhance our sequencing capabilities, such that ‘genomic surveillance’ may soon become the first line of investigation in all clinical research studies ongoing at our institution.” “Since the first published draft sequence of the human genome, our knowledge in genetics has exponentially increased,” says Dr. Ronald Cohn, co-director of the SickKids Centre for Genetic Medicine. “With the help of this new technology, we will be able to further deepen our understanding of the genetic basis of human disease and translate this directly into daily clinical practice. We have finally reached a point, where individualized medicine is not just a theoretical concept, but will become an integral part of clinical care and management.” The Ion Proton™ Sequencer is designed to sequence an entire human genome in a day for $1,000. Unlike traditional next generation systems, it relies on semiconductor chips to map human exomes and genomes, making it much faster and less expensive to analyze DNA at unprecedented throughput levels and generate accurate sequencing data. The system is based on the same technology as its predecessor, the Ion Personal Genome Machine (PGM™), which is designed for sequencing small genomes or sets of genes. Combined with Life Technologies’ AmpliSeq™ targeted sequencing technology, researchers can sequence panels of genes associated with disease on the PGM™ or exomes and genomes on the Ion Proton™ Sequencer in just a few hours.

Western Health’s regional director of laboratory services recognized by national body

Regional director of laboratory services with Western Health, Hedy Dalton Kenny is this year’s recipient of the A.R. Shearer Pride of the Profession award. The award was presented by the Canadian Society for Medical Laboratory Science during the opening Ceremonies at CSMLS’s recent conference LABCON 2012 in Gatineau, QC. “During her (25 year) career with

Western Health, Hedy has proven to be a dedicated technologist, manager and leader,” said Dr. Ken Jenkins, vice president of Medical Services. “She is committed to safety and quality in the lab and is persistent in its pursuit. Under Hedy’s leadership, Western Health received back to back accreditation from both Accreditation Canada and Ontario Laboratory Accreditation.” The CSMLS established the A.R. Shearer Pride of the Profession award to honour those that exemplify leadership in their profession. The award is named in honour of Archie Shearer, CSMLS executive director from 1961 to 1980. Shearer was an influential leader who was honoured with the Order of Canada for his contribution to Canadian health care during his time with the CSMLS. The A. R. Shearer Pride of the Profession award recognizes technologists who demonstrate professional pride through their leadership and commitment to excellence in the practice of medical laboratory science.

Beta amino acids for protein-based drugs Beta amino carbonyl structures are found in many pharmaceuticals. These structures are often associated with small synthetic proteins, or peptides, which are increasingly being considered as new pharmaceutical candidates. The challenge for peptide-based drugs has been that the body is well designed to break down proteins in the digestive system before they can make their way to the target. Dr. Andre Beauchemin of the University of Ottawa has been awarded $40,000 in Proof of Principle funding from GreenCentre to address this challenge. Dr. Beauchemin discovered a way to make synthetic beta amino acids with an “extra” carbon that can “fool” the digestive system into not tearing the proteins apart before they can take effect. Consequently, this makes it possible for pharmaceuticals based on this technology to be delivered orally, therefore increasing their potential. Beta amino carbonyl structures are found in many pharmaceuticals.

Dr. Andre Beauchemin

about Canada’s opportunity to truly make a difference in the world.” For the past 19 years, the Sanofi BioGENEius Challenge has given over 4,000 teen scientists access to university labs and academic mentors, promoting Canada’s biotech industry. This year, over 240 high school and CEGEP students from all over the country submitted 192 projects ranging from exploring prospective new drug treatments for Parkinson’s disease and multiple sclerosis to agriculture and the environment. To learn more about Song’s project, visit http://bit.ly/IrvD9I. Tam’s research can be found online at http://bit.ly/ Jw8mrq.

Hedy Dalton Kenny

scientific community believes in the potential of my research into tree particles as an eco-friendly alternative.” To enter the Boston International competition, Tam and Song participated in the 2012 Sanofi BioGENEius Challenge in Canada. “The Sanofi Group in Canada is thrilled to see that the potential of Canadian youth is being recognized at an international level,” said Mark Lievonen, president of Sanofi Pasteur Canada, in a prepared statement. “As the founding sponsor of the Sanofi BioGENEius Challenge Canada (SBCC) 19 years ago, we believe in the potential of our youth to develop the next big breakthrough in science. I am increasingly optimistic

These structures are often associated with small synthetic proteins, or peptides, which are increasingly being considered as new pharmaceutical candidates. The challenge for peptide-based drugs has been that the body is well designed to break down proteins in the digestive system before they can make their way to the target. These beta amino acids can be prepared at relatively low temperatures and at high yields, without expensive (and scarce) transition metal catalysts, and with low cost reagents, giving them several economic and environmental advantages.

July 2012 Laboratory Focus www.bioscienceworld.ca

News Continued from page 2 molecules, increasing their stability in the body and providing a higher degree of cell permeability. The platform adapts to any linear peptide input and has the power to generate libraries of compounds to facilitate drug discovery and development. This technology, discovered in Professor Andrei Yudin’s chemistry laboratory at the University of Toronto, will be developed with the collaboration of Professor Eric Marsault, specialized in medicinal chemistry at the Institut de Pharmacologie de Sherbrooke of Université de Sherbrooke. Together, they hope to demonstrate that Encycle’s foundational chemistry technology functions for a wide variety of peptides and that the platform can hit therapeutic targets of interest. “It costs over $1 billion to bring a new drug to market under the current high through-put pharmaceutical discovery model,” says Yudin. “Instead of randomly searching millions of compounds, Encycle’s platform will allow us to design a peptide molecule with small molecule properties, such as stability and cell permeability, while remaining more likely to interact with a targeted therapeutic area. In theory, this approach would save time, money and reduce overhead risk.” “The approach pioneered by Andrei

Yudin is remarkable for its efficiency in the synthesis of macrocycles, which are otherwise very difficult to reach and thus difficult to exploit in drug discovery,” says Marsault. “This collaboration will unlock the potential of this class and provide much-needed new classes of drug candidates able to mimic the natural structural elements of proteins.” The project has received $1 million in seed funding, largely through the Québec Consortium for Drug Discovery’s (CQDM) funding programs, and has attracted interest from four pharmaceutical companies. The joint provincial project is one of two pilot projects undertaken as part of the Ontario-Québec Life Sciences Corridor, which was announced at the 2011 Bio International Convention to foster existing strengths within the two provinces and increase innovation, productivity, investment and job creation. Based in part on Encycle’s success, and that of another undisclosed pilot project, CQDM and MaRS Innovation are partnering with the Ontario Centres of Excellence (OCE) and the Ontario Brain Institute (OBI) to launch the Québec/Ontario CQDM Funding Program, which will support similar collaborative research projects that seek new tools for biopharmaceutical research.

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promising new treatment for eBola infections

An Ebola virus. Photo: Frederick Murphy

Researchers at the Public Health Agency of Canada’s National Microbiology Laboratory (NML) have developed a new and easy-to-manufacture treatment for Ebola infection, one of the world’s deadliest diseases. The findings were published in the Science Translational Medicine journal. Zaire Ebola virus is one of the world’s most aggressive viruses. Up to 90 per cent of infections result in death within days of exposure. As there is no approved Ebola vaccine to prevent infection, there is an urgent need for a treatment to improve survival rates after exposure. Researchers say this new treatment

can be effective when administered up to 48 hours after infection. “Our researchers have seen first-hand the terrible effects of the Ebola virus on populations in Africa,” said Dr. Frank Plummer, chief science officer at the Public Health Agency of Canada. “This discovery should pave the way for the development of a new drug that has the potential to save many lives.” While Ebola does not naturally occur in Canada, there is always a small risk that it could be imported into Canada by an infected traveler. Having a safe and effective treatment option at the ready is important to protect Canadians from that risk. The NML is Canada’s leading public health infectious disease laboratory and the only facility in Canada that is permitted to study and work with live haemorrhagic fever viruses such as Ebola and other similarly highly infectious and deadly organisms.

Pfizer CaNaDa aNNouNCes NeuroPaThiC PaiN researCh awarD reCiPieNTs Funding from the Government of Canada is helping colleges across the country support business innovation at more than 300 companies. Minister of State for Science and Technology Gary Goodyear announced a $15.3 million investment through the Canada Foundation for Innovation (CFI) in support of 17 applied research projects at colleges, cégeps and polytechnics. The idea according to Minister Goodyear is Canadian companies specializing in everything from natural resources to information technology are turning to researchers at local colleges to grow their businesses and create high-quality jobs, because R&D and innovation happen at all levels— from the academic labs all the way to the shop floor. “Canada’s colleges, cégeps and polytechnics play an important role as we forge ahead with Canada’s innovation agenda,” he said. “Innovation needs to be a central focus—from the research laboratory to the production floor. It is the key to maintaining our country’s position as a global economic leader.” Minister Goodyear also announced

4/30/2012 1:21:13 PM

the recipients of the CFI’s new College-Industry Innovation Fund at George Brown College, including a project in green building technologies worth close to $1 million. The project will allow the college to provide students and small and medium-sized enterprises with increased opportunities to pursue applied research projects, boosting regional economic development and graduate job-readiness. “Canadian colleges are lending their expertise to local companies, helping them to compete and grow,” said Dr. Gilles G. Patry, president and CEO of the CFI. “The CFI is ensuring that colleges have in place specialized facilities to support their role as important hubs of economic growth and job creation.” “These investments provide colleges with access to the people, resources and tools they need to be at the forefront of innovation,” said Suzanne Fortier, president of NSERC. “The ultimate goal is to create sustainable partnerships that will help sharpen our innovative edge and have a positive impact on the bottom line of our country and industry.”

www.bioscienceworld.ca

The IRCM Foundation announces the appointment of Brian Gelfand to its board of directors. Gelfand is vicepresident of institutional relations and market operations at the Montréal Exchange and has previously worked as a consultant, an attorney, and a special advisor to the Secretary General at the International Organization of Securities Commissions. He is a member of the Bars of Quebec, New York and Massachusetts. Canada’s National Institute for Nanotechnology (NINT) has named Dr. Marie D’Iorio as its new executive director. Trained as a physicist, Dr. D’Iorio’s expertise

is in nano-electronics. She has been acting as NINT’s interim director general since last year. Dr. D’Iorio joined NRC in 1983, where she established the first very low temperature, high magnetic field laboratory in Canada to study low dimensional electron systems in semiconductor heterostructures. She served as director general of the National Research Council of Canada Institute for Microstructural Sciences from 2003 to 2011. She obtained a PhD in solid-state physics in 1982 from the University of Toronto, and then spent a year as a post-doctoral fellow at the IBM Zurich Research Laboratory in Switzerland. Molecular biologist Dr. John Capone has been named as new vice-president (research) of the University of Western Ontario. Currently serving his second term as McMaster University’s dean of science, Dr. Capone is recognized as one of Canada’s foremost molecular biologists. He begins his five-year term on October 1. In his

Laboratory Focus July 2012

aPPoiNTmeNTs pointed dean of McMaster’s Faculty of Science on July 1, 2005.

new position at Western, Dr Capone says he plans to build research capacity through faculty renewal, multi-disciplinary partnerships, embedding research at the undergraduate level and connecting research activity to educational excellence. He obtained a BSc in biochemistry from Western in 1978 and a PhD in biochemistry from McMaster University in 1983. From 1983 to 1986, he was a Medical Research Council of Canada centennial post-doctoral fellow at the Massachusetts Institute of Technology (MIT). He returned to McMaster as an assistant professor in the department of biochemistry in 1986 and was promoted to professor in 1995. He became chair of the department of biochemistry in 1997 and was appointed McMaster’s faculty of health sciences’ associate dean of research in 2000. Following a five-year term, he was ap-

Winnipeg-based biopharmaceutical company Cangene Corporation announces it has appointed Jeff Lamothe as its new chief financial officer commencing Aug. 1, 2012. Lamothe brings over 20 years business and financial experience to Cangene. He will be working out of Cangene’s Winnipeg headquarters, where he will oversee the hospital-based therapeutics company. Currently, Lamothe is CFO for Smith Carter Architects and Engineers Incorporated. Before that, he was president and CEO of Kitchen Craft Cabinetry, after occupying the position of VP Finance and CFO at that company. His past experiences include being CFO of Motor Coach Initiatives and working at James Richardson & Sons, Limited and Ernst & Young LLP. The Ontario Hospital Association (OHA) welcomes Pat Campbell as its new President and CEO. Campbell is an accomplished health care leader with a wide range of experience in rural, community and academic hospital environments. She joins the OHA from Echo: Improving Women’s Health In Ontario, an agency of the Ministry of Health and Long-Term Care. Before that, Pat served as

president and CEO of Grey Bruce Health Services and Women’s College Hospital in Toronto. BIOTECanada announces the appointment of Andrew Casey as the association’s new president and CEO starting July 30, 2012. Casey joins the biotech industry after spending almost 20 years in senior roles in the national trade association business. Currently, Casey is the vice president of public affairs and international trade with the Forest Products Association of Canada (FPAC). Prior to joining the FPAC, Casey was assistant vice president, government relations, for the Canadian Life and Health Insurance Association. The announcement was made by Brad Thompson, the chairman of BIOTECanada, who welcomed Casey to the industry and to the association.

Sue Paish has been named the new CEO for LifeLabs. She joins LifeLabs from Pharmasave Drugs. Prior to joining Pharmasave, Paish was the managing partner of Fasken Martineau DuMoulin in B.C. She sits on a number of corporate and community boards including the Michael Smith Foundation for Health Research, the Rick Hansen Foundation and the CORIX Group of Companies. She begins in her new role as CEO of LifeLabs on July 1. David Gauthier succeeds Wilf Keller as president of Genome Prairie. Gauthier is a private consultant specializing in investment and technology commercialization. He has served as director of business development with Performance Plants Inc., vice-president of Foragen Technologies Management Inc., regional director of the National Research Council’s Canada Industrial Research Assistance Program and most recently CEO of the Entrepreneurial Foundation of Saskatchewan. Stepping down as Genome Prairie’s president allows Keller to make his current role with Ag-West Bio a full-time position. Keller had held the dual roles since April 2010, when a management agreement was struck between the two organizations

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July 2012 Laboratory Focus www.bioscienceworld.ca

Pharma Notes

Dalton Pharma Services (Toronto, ON) has received GLP certification from The Standards Council of Canada. SCC accreditation is recognized around the globe, while in Canada, SCC administers the OECD GLP initiative. OECD (Organization for Economic Development and Co-operation) has 34 member countries, including Canada, Germany, Japan, and the US. Ildiko Riss, director of quality at Dalton, noted “The auditing procedure of SCC was very rigorous. The GLP certificate issued as a result of the inspection and audits covers the field of pharmaceuticals and the following areas: analytical and clinical chemistry testing;

pharmacokinetic testing; and physical-chemical testing. Dalton’s certificate can be viewed on the SCC website (www.scc.ca).” Peter Pekos, president and CEO further commented “The confirmation of Dalton’s GLP compliance is evidence of this commitment to quality. I am very proud of the dedication and accomplishments of our Quality Group.” This year marks Dalton’s 25th year in business. The Centre for Drug Research and Development (CDRD) (Vancouver, BC) , its commercial arm, CDRD Ventures Inc. (CVI), and GlaxoSmithKline (GSK) in Canada have entered into

a strategic collaboration that will provide new financial resources as well as leading drug development expertise to be applied to the development and commercialization of health research conducted in Canadian research institutions. Under the collaboration, GSK will provide project-based funding to support the commercialization of health research. The first such vehicle is a “GSKCDRD Innovation Fund” within CDRD to support certain early-stage projects to be conducted by CDRD and carried out in collaboration with academic investigators at CDRD’s affiliated institutions and hospitalbased research centres. A

THE 13th INTERNATIONAL CONFERENCE ON SYSTEMS BIOLOGY

AUGUST 19–23, 2012 TORONTO, CANADA Plenary Speakers: Jurg Bahler, University College London Thijn Brummelkamp, Netherlands Cancer Institute David Botstein, Princeton University Michael Boutros, DKFZ Heidelberg George Church, Harvard Medical School Anne-Claude Gavin, EMBL Heidelberg Tim Hughes, University of Toronto Ben Lehner, EMBL-CRG Systems Biology Unit, Barcelona Norbert Perrimon, Harvard Medical School Pam Silver, Harvard University Mike Snyder, Stanford University Eran Segal, Weizmann Institute

Organizers: Brenda Andrews, Gary Bader, Charlie Boone, Cynthia Colby, Roland Eils, Anne-Claude Gavin, Stefan Hohmann, Michael Hucka, Tim Hughes, Roy Kishony, Hiroaki Kitano, Edda Klipp, Stephen Michnick, Corey Nislow, Fritz Roth, Sachdev Sidhu, Michael Snyder Sponsor:

joint innovation committee comprised of both GSK and CDRD representatives as well as external reviewers will review and select the projects to be supported from this fund. The second fund is a joint venture with CVI that will see additional funds invested in advancing projects further towards commercialization. Under this collaborative relationship, CVI will then conduct (or manage the conduct of) multiple selected research projects to be partially funded by GSK with the goal of commercializing the technology and resulting intellectual property. Valeant Pharmaceuticals International, Inc. (Montréal) has agreed to acquire OraPharma (Horsham, PA), a specialty oral health company that develops and commercializes products that improve and maintain oral health, from Water Street Healthcare Partners, a private equity firm focused exclusively on the health care industry. Total consideration is approximately $312 million and up to $114 million in potential contingent payments based on certain milestones, including revenue targets. OraPharma’s lead product is Arestin, a locally administered antibiotic for the treatment of periodontitis that utilizes an advanced controlled-release delivery system and is indicated for use in conjunction with scaling and root planing for the treatment of adult periodontitis. The NCIC Clinical Trials Group at Queen’s University in Kingston, ON has entered into an agreement with Oncolytics Biotech Inc. (Calgary, AB) to sponsor and conduct a randomized Phase 2 study of REOLYSIN® in patients with advanced or metastatic breast cancer. The study will be an open-label, randomized, non-blinded, Phase 2 clinical study of REOLYSIN given in combination with paclitaxel versus paclitaxel alone. Approximately 50 response-evaluable patients will be enrolled in each arm, after a six to nine patient safety run-in.

Stellar Pharmaceuticals Inc. (London, ON) has acquired the Canadian rights for Collatamp G® from Theramed Corporation (Mississauga, ON). Collatamp G® is approved in over 50 countries for the local haemostasis of capillary, parenchymatous and seeping haemorrhages in areas with a high risk of infection and has been shown to reduce postoperative infections across a range of surgical disciplines, including a reduction of 53 per cent in a large randomized controlled study in cardiac surgery. Collatamp G® was approved by Health Canada on Aug 1, 2007 and launched in Canada in 2008. Amorfix Life Sciences Ltd. (Mississauga, ON) has signed an agreement granting an exclusive worldwide license for its preclinical Alzheimer’s disease diagnostic test, the Amorfix Aggregated Abeta Assay (the A4) to JSW Lifesciences GmbH, a contract research organization specializing in Alzheimer’s disease and other neurodegenerative disorders. Under the terms of the agreement, JSW will market and perform the A4 assay as a service in the area of preclinical Alzheimer’s disease studies. The agreement includes a commitment to minimum annual sales and Amorfix will receive a percentage of net sales. Nuvo Research Inc. (Mississauga, ON) and Paladin Labs Inc. (St. Laurent, QC) have entered into a license and supply agreement granting Paladin exclusive Canadian rights to market and sell Synera®, a topical patch that combines lidocaine, tetracaine and heat. Under the terms of the agreement, Nuvo will receive a double digit royalty on net sales of Synera in Canada and will supply Synera to Paladin. In addition, Paladin has agreed to loan Nuvo $8 million in two equal tranches of $4 million each (the first drawn on closing and the second to be drawn on the achievement of milestones).

www.bioscienceworld.ca

Laboratory Focus July 2012

feaTure

By: roBerT e. CamPBell, uNiversiTy of alBerTa

New Bioanalytical

Tools and Devices Chemistry leads the way

iNTRoDuCTioN Three university of Alberta chemists are developing innovative and imaginative bioanalytical techniques aimed at tackling the burden and suffering caused by infectious diseases in the developing world. Bioanalytical chemists are accustomed to having a plethora of convenient consumables and sophisticated instrumentation at their disposal. In the absence of this infrastructure it is hard to imagine how one might be able to achieve the primary goals of bioanalytical chemistry, the detection of small molecules or macromolecules in biological systems. Unfortunately, many researchers in resource-limited developing countries, or field workers in remote locations far from modern conveniences, are unable to take advantage of modern bioanalytical techniques due to a lack of infrastructure. Compounding the misfortune of this situation is that these same researchers are those who have the greatest need for rapid and accurate bioanalytical detection methods, typically to diagnose diseases such as tuberculosis or malaria, which are a disproportionate burden on the developing world. New bioanalytical techniques designed from the bottom up to address the needs of researchers in resource-limited environments could help level the playing field and facilitate point-of-care diagnosis in

the developing world. Motivated by this shared goal, Dr. Julianne Gibbs-Davis, Dr. Ratmir Derda, and Dr. Michael Serpe in the department of chemistry are independently taking three very different approaches towards the development of new bioanalytical tools and devices. Each of these researchers is effectively leveraging the cutting edge infrastructure of a world class research institution to lower the financial threshold limiting access to modern bioanalytical chemistry.

Move over PCR! The polymerase chain reaction (PCR) is the method of choice for detecting low concentrations of DNA and is routinely used for the detection and diagnosis of viral and bacterial diseases. However, PCR requires the use of thermal cycler instrumentation that may not always be available in resource limited laboratories or sites that lack reliable access to electricity. In such situations, it would be preferable to use an instrumentfree method to amplify the concentration of DNA in a dilute sample until reaching a concentration where it

“By keeping the assay simple and minimizing the complexity of the technology, we are hopeful that this will be more easily adopted in resourcelimited settings.” — Dr. Julianne Gibbs-Davis

amplification using figure 1 DNA cross-catalytic ligation.

July 2012 Laboratory Focus www.bioscienceworld.ca

Feature Figure 2

“The first functional prototype of the phage-based diagnostic could be ready to showcase after a few months.”

An origami Petri dish for assessing the growth of coloured bacteria.

— Dr. Ratmir Derda

tection, for diagnosis of tuberculosis and multi-drug resistant tuberculosis. As Gibbs-Davis explains, “By keeping the assay simple and minimizing the complexity of the technology, we are hopeful that this will be more easily adopted in resource-limited settings.” With continued optimization, she expects that it will be possible to get the reagent costs under a few dollars per test, making her method cost competitive with traditional PCR-based methods.

Origami Petri dish

can be easily detected using common fluorescence-based methods. One potential application would be the inexpensive diagnosis of tuberculosis by detection of characteristic DNA sequences from the causative agent, Mycobacterium tuberculosis. In an effort to reach this goal, Gibbs-Davis turned to an idea that she had been eager to try since her days as a graduate student at Northwestern University. Briefly, GibbsDavis envisioned a system in which one strand of DNA would template the enzymatic ligation of two partially complementary fragments. Normally, such a reaction would produce a double stranded DNA duplex that would be substantially more stable than the free single strand DNA pieces and thus represent the end of the reaction process. Gibbs-Davis’s ‘eureka’ moment of insight came when she realized that she could engineer the system such that the DNA duplex was destabilized enough that it would occasionally dissociate to form the single stranded pieces (Figure 1). This dissociation frees the template strand to engage in another cycle of ligation, essentially acting as a catalyst for the ligation reaction of the two complementary fragments. In their initial versions of this design, Gibbs-Davis and her coworkers succeeded in coaxing a template strand

into performing 18 cycles of ligation, or turnovers, in 20 hours, a substantial improvement over previously reported ligation-based amplifications under similar conditions.1 The first generation of Gibbs-Davis’s amplification system detects small DNA fragments that are complementary to the template sequence. To detect a longer and intact target DNA sequence, Gibbs-Davis devised a cross-catalytic ligation cycle in which a target DNA sequence, necessarily composed of unmodified DNA, ligates two partially complementary strands to make a new destabilizing template that can participate in the catalytic ligation of additional fragments. In this way a piece of target DNA acts as a trigger for initiating the catalytic cycle of DNA amplification. With the latest version of her cross-catalytic ligation system, Gibbs-Davis can now get thousands of copies of template molecule in just a few hours of incubation. While this is still not approaching the billions of copies that can be obtained from PCR amplification, it is already practical for some applications, and further improvements are likely. Gibbs-Davis plans to incorporate this amplification system into an inexpensive kit, which will include a sample holder with a built-in light emitting diode for fluorescence de-

Compared to the traditional animal shapes and intricate geometric forms produced by the Japanese art of origami, a Petri dish seems to be a particularly unlikely structure to create from a sheet of paper. However, this has not stopped Dr. Ratmir Derda from developing a Petri dish replacement from a sheet of paper that has been cleverly folded and manipulated to create a sterile environment for cell growth. Such devices could enable researchers in the developing world to carry out cell-based bioanalytical assays for research purposes or for detection of disease-causing agents. To achieve the goal of creating a paper-based cell culture system, Derda has abandoned the paradigm of growing bacteria in an open dish (i.e. a Petri dish) with a loose fitting lid. Rather, he has designed a selfcontained system in which bacteria grow inside a sterile paper envelope that retains liquid growth media, yet is permeable to oxygen (Figure 2). This growth chamber has a transparent window such that a researcher can easily visualize the growth of the bacteria inside. A particularly effective strategy for facilitating the visualization of bacteria growth in the chamber is to introduce a gene for a brilliantly coloured red fluorescent protein. In this way, the amount of bacterial growth can be qualitatively assessed by eye, or quantitatively assessed using a camera phone and a custom software application. In tackling the problem of how to most effectively facilitate cell culture techniques in resource-limited environments, Derda endeavoured to find a convenient testing ground. His inspiration was to turn to Canadian

high school classrooms, which, relative to chemistry labs at the University of Alberta, represent a resourcelimited environment that could approximate the level of infrastructure one could expect to find in a lab in the developing world. Accordingly, Derda has been working closely with high school students to optimize the procedure for mass production of his paper-based Petri dishes. With the support of a Canada’s Rising Stars in Global Health grant from Grand Challenges Canada, Derda is exploring the combined use of paperbased petri dishes and engineered bacteriophage for biosensing applications. One of his primary goals is to use this system for detecting tuberculosis-specific antibodies in serum and he says that “the first functional prototype of the phage-based diagnostic could be ready to showcase after a few months.” Over the longer term, he foresees application to a broader array of bioanalytical applications, such as rapid and inexpensive diagnostics for HIV or malaria. Derda has also used a substantial portion of his Rising Star award to organize a workshop in Kenya that will bring together like-minded researchers who are working towards the development of diagnostic devices for use in resource-limited environments. Derda hopes that, by holding the conference in Africa, he will have the opportunity to give a number of North American researchers first hand experience of the actual level of resources available in these ‘low resource’ environments. When asked about his own recent trip to Kenya, Derda replied, “Many laboratories I saw were ghost labs filled with equipment that cannot be used because the lab does not have consumables.” Apparently, programs that redistribute surplus equipment from well-funded labs have been very successful, but the problem is that much of this equipment relies on manufactured consumables that simply are not available. The solution, Derda says, is the concept of “on-site production.” Specifically, if researchers in developing countries could produce low-cost alternatives to common laboratory consumables they would be in a much better position to take advantage of the instrumentation re-

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July 2012

feaTure figure 3

“...the polymer can be made such that it interacts with, and binds to, biomolecules that are indicative of disease or infection.”

Colour changing etalons for use in bioanalytical applications.

— Dr. Michael Serpe already demonstrated that this principle can be applied to glucose sensing3 and, with the support of a Canada’s Rising Stars in Global Health grant from Grand Challenges Canada, is now working to extend the approach to a wider range of bioanalytical target molecules. Short-term goals include the detection of biomarkers for malaria and tuberculosis, while longer-term goals involve extending this technology to the detection of biomarkers for the early stages of Alzheimer’s disease. sources that are available to them.

Technicolor bioanalysis While not every lab in the developing world can afford a visible wavelength spectrophotometer, most humans have a built-in equivalent: their colour vision. The key to taking advantage of human colour perception for bioanalysis is having a robust method for converting the concentration of a target analyte into a visible colour change. Inspired by the brilliant blue wings of a butterfly, the Serpe group is now developing devices that achieve this goal. The surface of the Morpho didius butterfly wing is covered in small ridges that have just the right spacing to cause constructive interference of blue light, while other wavelengths are removed by destructive interference. In principle, if the butterfly was able to change the spacing of the ridges on its wings it could change its colour to green or red by causing constructive interference of the appropriate wavelengths of visible light. Although butterfly wings do not have this capability, the Serpe group is now building devices that are artificial mimics of the butterfly wing, yet are capable of changing their colour in response to a biomolecular target analyte.2 To create these colourchanging biosensors, Serpe

has turned to etalons: a sandwich structure composed of two reflective gold layers (the bread) on either side of a layer of microgel particles (the filling) (Figure 3). At a certain thickness of filling, the device appears a certain color due to the constructive interference of particular wavelengths of light. However, when the particles are induced to change their size, the thickness of the

filling changes and the colour of the device undergoes a corresponding change. Serpe explains that “...the polymer can be made such that it interacts with, and binds to, biomolecules that are indicative of disease or infection. Hence, the device’s colour change can be coupled to the presence of a specific biomarker for disease, and used as a diagnostic device.” Serpe has

Canadian chemistry leads the way This article has highlighted three exciting new approaches towards enabling robust bioanalytical detection methods that do not require the cutting edge infrastructure

and modern conveniences that have helped Canada become a world leader in bioanalytical research. With the seemingly unlimited resource of researcher ingenuity, and the continued generous support of funding agencies such as Grand Challenges Canada, Canadian researchers are well positioned to play a key role in the uplifting of nations who do not share our advantages.

References 1. Kausar, A.; McKay, R.D.; Lam, J.; Bhogal, R.S.; Tang, A.Y.; Gibbs-Davis, J.M., Angew. Chem. Int. Ed. Engl. 2011, 50(38), 8922-8926. 2. Hu, L.; Serpe, M.J., Polymers 2012, 4(1), 134-149. 3. Sorrell, C.D.; Serpe, M.J., Anal. Bioanal. Chem. 2012, 402(7), 2385-2393.

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Feature

B y: Alex P ower , Vincent Martin, and Mads Kæ rn

By: Sergey N Krylov

Direct Quantitative Analysis

of Multiple miRNAs (DQAMmiR) – a New Tool for Personalized Cancer Medicine Cancer is a heterogeneous disease - not only do different types of cancer differ from each other, but individual cancers of the same type also vary from patient to patient. Therefore, the classical “one-sizefits-all” approach in cancer treatment is not efficient. The base paradigm of personalized cancer medicine is that patient-specific characteristics of cancer must be used in choosing a therapy to make it efficient. Molecular markers in tumours are the most reliable means of characterizing individual cancers and microRNA (miRNA) is, in turn, among the most promising types of molecular markers. MiRNAs are 18 to 25-nucleotidelong strands of RNA that play an important role in regulation of cellular processes. Abnormal production of miRNA in cells was implicated in cancer suggesting that miRNA can be used for cancer diagnosis and therapy guidance. Tumourigenesis typically leads to altered levels of several miRNAs, for example, abnormal production of two miRNAs was linked to colorectal cancer in humans. Accurate molecular characterization of individual cancers requires identification of miRNA signatures of cancer, which are per se quantitative levels of multiple miRNAs. Hundreds of candidate miRNA markers of cancer have been already identified by comparing cancerous and normal tissues with microarrays which allow profiling thousands of miRNAs in a semiquantitative fashion. Despite these efforts, currently there is no single

miRNA signature validated for cancer prognosis and therapy guidance. Validation of miRNA signatures is the main obstacle to using miRNA in personalized cancer medicine, and the lack of a suitable validation technology for miRNA signatures is arguably the major challenge. To validate a miRNA signature, one needs to accurately measure levels of multiple miRNAs in order to then establish the correlation of these levels with clinical outcomes of cancer for a statistically significant population of patients (as many as tens of thousands of patients). Accordingly,

a suitable validation technology must be: (i) quantitative, (ii) highly-sensitive, (iii) rugged, and (iv) capable of simultaneously analyzing multiple miRNAs. Quantitation is required since cancer sub-types may differ from each other in varying quantities of miRNAs in the molecular signature. High sensitivity is required to allow analyses of very small tissues samples; for instance, those from fine-needle aspiration biopsies or single cells. Ruggedness is needed to ensure that data obtained in different labs with different instruments are comparable (the effort of many

labs is needed to process a very large number of samples required for validation). Finally, the simultaneous analysis of multiple miRNAs is needed to generate uniform signatures as well as reduce cost of analysis, thus, making validation of miRNA signatures feasible. Most available methods of miRNA detection are indirect (e.g. quantitative reverse-transcriptase polymerase chain reaction, microarrays, surface plasmon resonance, next generation sequencing, etc.); they require chemical or enzymatic modifications of miRNA prior to the

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Laboratory Focus July 2012

feaTure analysis. Modifications make the analysis cumbersome and lead to reduced accuracy due to different efficiencies of modifications for different miRNAs. All indirect methods do not satisfy more than one of the four conditions specified above for a suitable validation technology. A few direct methods, which do not require any modification of the target miRNA, are available: Northern blotting, signal amplifying ribozymes, in situ hybridization, bioluminescence detection, and twoprobe single-molecule fluorescence. However, they have limitations – the first three are only semi-quantitative when used for multiple miRNAs while the latter two can hardly be used for multiple miRNAs. Direct Quantitative Analysis of Multiple miRNAs (DQAMmiR) that has a potential to satisfy all four requirements has been recently developed by a group of researchers from York University upon a classical hybridization approach.1 In this approach, hybridization probes fluorescently labeled for detection and taken in excess to miRNAs, bind miRNAs sequence-specifically, and unreacted probes are separated from the probe-target hybrids by capillary electrophoresis and detected with laser-induced fluorescence. The challenging problems of electrophoretic separation of (i) the unreacted probes (single strand DNA) from the probemiRNA hybrids and (ii) the hybrids from each other were solved through the combined use of single strand DNA binding (SSB) protein in the electrophoresis run buffer, and drag tags on the probes which change the probe electrophoresis mobility. Figure 1 schematically illustrates the concept of DQAMmiR. In the hybridization step, the excess of the probes of concentrations [P]0,i is mixed with the miRNAs to be analyzed, which leads to all miRNAs’ being hybridized but with some probes left unreacted. A short plug of the hybridization mixture is introduced into a capillary filled with an SSB-containing run buffer. Due to its unique binding selectivity, SSB binds all unreacted ssDNA probes but does not bind the double stranded miRNA-DNA hybrid.

When an electric field is applied to the ends of the capillary, the SSB-bound probes move faster than all the hybrids (SSB serves the role of the propellant). The miRNADNA hybrids have similar lengths and cannot be easily separated by electrophoresis;

however, different drag tags on the probes make different hybrids move with different velocities. SSB-bound probes will move with similar velocities if the drag tags are small with respect to SSB. In such a case, a fluorescent detector at the end of the capillary

generates separate signals for the hybrids and a cumulative signal (one peak or multiple peaks) for the unreacted probes. The amounts (or concentrations) of miRNAs in the sample are finally calculated with a simple mathematical formula that uses the inte-

grated signals which correspond to peak areas in the graph: AH,i for the hybrids and AP for a sum of all unreacted probes. DQAMmiR is quantitative and accurate. It does not involve amplification or chemical/enzymatic modifications

July 2012 Laboratory Focus www.bioscienceworld.ca

Feature

that could affect accuracy of quantitation. The dynamic range of DQAMmiR is easily adjusted by changing the concentrations of the hybridization probes. Not only is the DQAMmiR quantitative but it also allows one to determine absolute amounts of miRNAs in the sample by using the unreacted probes as a reference standard. This internal reference standard makes DQAMmiR calibrationfree which is the method’s unique and powerful feature. DQAMmiR can, thus, be used for distinguishing miRNA signatures that differ in miRNA amounts. With regards to DQAMmiR sensitivity, commercial capillary electrophoresis equipment can only facilitate detection limits of approximately 300,000 copies of miRNA in a consumed sample. With this relatively high detection limit DQAMmiR would be inapplicable to measuring miRNAs in fine-needle aspiration biopsies, which became standard in diagnosis of cancer and other diseases. Indeed, as few as 1,000 copies of miRNA may be present in a single cell, and fineneedle biopsies typically contain in the order of 100 cells. The total number of miRNA copies in such a small sample would be less than 100,000, which is below the DQAMmiR detection limit possible with commercially available CE instrumentation. To solve this problem, the researchers from York University collaborated with Prof. Gradinaru’s group at the University of Toronto, Mississauga, to develop ultra-sensitive capillary electrophoresis (CE) with confo-

Figure 1

Separaation by SSB‐mediated CE

Hybridization

DNA probes

Electric field Injection to o CE

miRNA

ssDNA biinding protein (SSB) (

Detection Flu uorescen nce intensityy

While the ruggedness of DQAMmiR has not yet been confirmed experimentally, one can expect the method to be rugged. DQAMmiR is a calibration-free method that can determine absolute amounts of miRNA. The results do not depend on an instrument used provided that it has a suitable sensitivity.

Detector

Quantitation of miRNA N

[miRNA]i 

AH,i

A H, i  [P]0, i N

A i

H, i

 AP

Migration time Schematic illustration of the direct quantitative analysis of multiple miRNAs (DQAMmiR) by CE with fluorescence detection. miRNAs and their complimentary ssDNA probes are shown as short lines of the same color, drag tags are shown as parachutes, a fluorescent label is shown as small green circles and ssDNA-binding protein (SSB) is shown as large black circles.

cal time-resolved fluorescence (CTRF) detection through an embedded capillary interface (ECI).2 CE-CTRF-ECI decreased the limit of detection to 1,000 copies of miRNA, and further improvement can be achieved with more advanced prototypes of CE-CTRF-ECI instrumentation. Importantly, instrumentation for CE-CTRF-ECI is not only sensitive but also robust; it does not include sophisticated custom-made components that are easy to align and keep alignment. DQAMmiR performed with CE-CTRF-ECI can, thus, be used for analysis of miRNAs in a small number of cells and even in single cells. While the ruggedness of DQAMmiR has not yet been confirmed experimentally, one can expect the method to be rugged. DQAMmiR is a calibration-free method that can determine absolute amounts of miRNA. The results do not depend on an instrument used provided that it has a suitable sensitivity. Moreover, DQAMmiR’s performance is not significantly dependent on the sample matrix and even a crude cell lysate can be sampled for the analysis. This suggests that the method has minimal susceptibility to variations in sample preparation procedures, thus allowing different laboratories and operators to

obtain comparable data. This in turn will facilitate screening large populations of patients required for detailed profiling of cancer sub-types. DQAMmiR is capable of simultaneously analysing multiple miRNAs. The number of miRNAs that can be run together is limited by the number of hybrid peaks that can be baseline-separated in the time window between a peak of miRNA hybrid with untagged probe and a peak of SSB-bound unreacted probe (see Fig. 1). The current estimate suggests that simultaneous analysis of 10 different miRNAs should be easily achievable. This number can be further increased by a factor of two or three if two or three different fluorophores are used as labels along with two or three spectral channels in the instrument. While being potentially suitable for validation of miRNA signatures of cancer, DQAMmiR still requires significant research effort to become a practical tool in personalized cancer medicine. A generic approach to the design of multiple hybridization probes with different drag tags is still to be developed. For DQAMmiR to be used by multiple groups, commercial instrumentation for CE-CTRF-ECI must be available. Finally, DQAMmiR still need to be rigorously validated before it can

be widely used in laboratory and clinical settings. The proof of principle for DQAMmiR and encouraging initial results certainly stimulate the effort towards making it a practical research and clinical tool.

References: 1. Wegman, D.W.; Krylov, S.N. Direct quantitative analysis of multiple miRNAs (DQAMmiR). Angewandte Chemie International Edition 2011, 50, 10335 –10339. 2. Dodgson, B.J.; Mazouchi, A.; Wegman, D.W.; Gradinaru, C.C.; Krylov, S.N. Detection of a thousand copies of miRNA without enrichment or modification. Submitted to Analytical Chemistry on June 5, 2012.

Dr. Sergey Krylov was educated at Moscow State University and is now Professor of Chemistry, Canada Research Chair in Bioanalytical Chemistry, and Director of Centre for Research on Biomolecular Interactions at York University. He is a winner of a number of awards including the 2007 W.A.E. McBryde Medal of the Canadian Society for Chemistry for a significant achievement in pure or applied analytical chemistry.

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Laboratory Focus July 2012 By yuJie zhu aND Jose m. moraN-miraBal

feaTure

Micro and

nanostructured materials for the study and monitoring of biomolecular interactions The ability to control, monitor and study biomolecular interactions, such as recognition, binding, catalysis and signal transduction, is critical not only for understanding fundamental cell biology but also for the design of efficient high-throughput biosensing and diagnostic methods. Micro and nanostructured surfaces have been targeted as powerful tools to investigate biomolecular interactions because they yield high-density arrays of biomaterials at biologically relevant scales. On one hand, they can produce motifs that can be used as simplified surrogates for the complex cellular microenvironment. This has enabled the in vitro study of targeted biomolecular interactions, which would be hard or impossible to study in vivo. On the other hand, the demand for efficient detection of proteomic and genomic biomarkers, as well as the need for rapid detection of pathogenic threats has placed great emphasis on the production of biofunctional micro and nanostructured materials that act as recognition elements in high-throughput biosensing platforms. Such materials enable the specific binding and sensing of target biomolecules in complex mixtures with high sensitivity and low noise. Because of the potential applications in cell biology and in biosensing and diagnostic tools, much research has focused on the ability to create bioactive micro and nanostructured surfaces. To date, numerous techniques have been developed to accomplish this goal including: self-assembly, micro and nanomachining, quill-pin spotting, dip-pen nanolithography, inkjet printing, microcontact printing, polymer stencil lift-off, electrospinning, and micro-fluidic networks, among others. Our laboratory focuses on the development of techniques to produce surfaces that enable the immobilization of biomaterials in controlled micro to nanoscale patterns, and their application for the study of targeted biomolecular interactions. Below, we detail two specific techniques that we currently employ, and discuss their potential applications in the fields of cellular and molecular biology, and in

the development of biosensors and point of care devices.

Polymer stencil lift-off for biomaterial micropatterning Biomaterial micropatterning refers to the host of techniques used to deposit biomaterials on solid surfaces with controlled feature sizes of micron to nanometer dimensions. Micropatterning is a powerful and rapid means for producing arrays of biomaterials for biosensing assays and for the study of interactions between the patterned materials and target analytes. A number of biocompatible techniques have been developed aimed at placing biological ligands at welldefined locations on substrates, with the most widely used being microcontact printing (µCP). Although strategies for DNA and protein patterning are relatively mature, supported lipid bilayer (SLB) and cellular patterning are more challenging because they require the materials to remain in ‘nev-

er-dry’ condition and cannot be easily achieved by any of the currently available techniques. An additional shortcoming of current techniques lies in the ability to deposit a multiplicity of biomaterials to form intricate patterns. Our research group is working on the development of new approaches to overcome these challenges. Among the techniques developed for biomaterial micropatterning, polymer stencil lift-off (PSLO) is one of the most robust and is utilized in our lab due to its advantages in terms of biocompatibility, pattern transfer fidelity, and its applicability to patterning under aqueous conditions. PSLO is a relatively new patterning method that enables the formation of arrays of micro to nanometre-sized biomaterial domains over large surface areas. In this approach, Parylene C (dichloro-[2,2]paracyclophane) is the most commonly used polymer because it produces chemically inert, pinhole-free, thin polymer films. Furthermore, these

films do not swell and can be mechanically peeled off from the surface under aqueous conditions, which makes them ideal templates for patterning biomaterials that need to be kept in hydrated environments (e.g. lipid membranes and cells). The principle behind PSLO micropatterning is illustrated in Figure 1. First, a thin conformal polymer film is deposited onto the solid surface through chemical vapour deposition. A photoresist layer is then coated on the polymer and patterned using photolithography. With the patterned photoresist as a mask, the sample is subjected to a controlled oxygen plasma reactive ion etch that removes the polymer and exposes the underlying substrate surface, effectively creating a polymer stencil. The biomaterials of interest can then be applied and bound onto the exposed substrate, while the polymer stencil keeps all the covered areas protected. After the excess biomaterials are washed away, the stencil can be me-

July 2012 Laboratory Focus www.bioscienceworld.ca

feature chanically easily peeled off due to its weak adhesion to the substrate, leaving the biomaterial micropattern on the surface. Currently, research in our lab focuses on supported lipid bilayer (SLB) micropatterning through the PSLO technique. SLBs are interesting model systems because they preserve many of the distinctive properties of cell membranes, such as fluidity and functionality, while at the same time allowing control of the membrane composition and ease of access for their study through quantitative analytical techniques. Thus, SLBs have become increasingly popular and offer opportunities for the study of lipid-lipid, ligand-receptor1, and cellmembrane2,3 interactions, as well as for the production of membrane based biosensors.4–6 SLB micropatterning provides the additional capabilities of controlling the size of the SLB domains and of producing SLB arrays of heterogeneous compositions. Furthermore, the combination of SLB micropatterning with microfluidic devices provides a powerful tool for studying biomolecule interactions with low sample consumption and high throughput. With the PSLO technique, our group is able to produce micron and sub-micron sized patterns of SLBs with homogenous and heterogeneous lipid compositions under aqueous conditions (Figure 1). We use micropatterned SLB arrays as models for the study of the effect of domain size and environmental factors (e.g. temperature, cholesterol concentration, ionic concentration) on the behavior of lipid mixtures. Through these simplified model systems we aim at understanding more complex systems such as the plasma membrane.

Electrospinning of Functional Nanofibers Developed over 70 years ago and popularized by Reneker in the 1990’s,

Figure 1

The polymer stencil lift-off technique produces well-defined, high quality micron and sub-micron supported lipid bilayer patterns. Top: Schematic description of the PSLO technique and illustration of the lift-off process. Bottom: sample images of line (3 μm wide) and square (10 μm wide) patterns formed using PSLO of fluorescently labeled SLBs with heterogeneous lipid mixture compositions.

electrospinning is another powerful technique for generating micro and nanostructured materials. In recent years, there has been increased interest in the study of electrospun nanofibers due to the surge in the demand for nanostructured materials. With electrospinning, not only a variety of polymers, but also biomolecules, nanoparticles, and even cells embedded in a carrier polymer, can be used to produce nanofibers with different physical, chemical and biological properties. In particular, fibers made of functional polymers

can possess unique mechanical7, electrical8, electrochemical9, magnetic10, and optical properties.11 With easily tailored properties and functionalities, electrospun nanofibers have promising applications in drug delivery, energy storage, tissue engineering, sensing and lab-on-a-chip applications. One of the most attractive features of the production of functional nanofibers through electrospinning is that can be done on the bench top in a very simple manner. In a typical electrospinning setup, such as the one implemented in our laboratory, a droplet

Figure 2

Electrospinnning is a simple and low-cost technique for producing nanostructured materials. Left: Schematic depiction of a standard electrospinning setup. Right: Fluorescence images of photoluminescent nanofibers produced from polymer solutions doped with different fluorogenic molecules.

of viscous solution is applied onto a sharp conductive tip and high electrical potential is applied between the tip and a static or rotating grounded collector. As a result of molecular ionization, charge redistribution and the external electric field, the droplet deforms until the electrostatic forces acting on the charged solution overcome surface tension. At this critical point a Taylor cone forms and a continuous jet is extracted from the solution. As the jet travels through the air from the tip to the collector, it is thinned out due to solvent evaporation, stretching and bending forces, which results in the formation of very narrow solid nanofibers that can be deposited on a substrate affixed to the collector. The electrospinning process is mainly affected by the material properties of the polymer solution (molecular weight and polydispersity, viscosity, surface tension, and conductivity of the solution) and process parameters (amount of solution applied, electric voltage, distance between the tip and the collector, and motion of collector). With proper control of the solution viscosity, the voltage applied and the distance between the tip and the collector substrate, fibers with diameters in the sub-micron range can be easily produced via electrospining. Our current research explores bench top techniques to fabricate mi-

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Laboratory Focus July 2012

crostructured conductive tips that can be integrated with continuous flow microfluidic systems for the electrospinning of conductive polymer nanofibers. Our conductive tips are typically fabricated by directly cutting polymer films, which are subsequently coated with a conducting layer. This approach allows us to test different shapes and sizes and tailor the electrospinning tip to the particular polymer blend that we wish to electrospin. Carrier polymers that we use for electrospinning include polystyrene (PS), poly-ethylene oxide (PEO), and poly-vinyl alcohol (PVA) dissolved in different polar and non-polar solvents such as water, ethanol, chloroform, and toluene, among others. The ability to pair the electrospinning tip morphology with a variety of polymer and solvent combinations allows us to routinely produce fibers with diameters smaller than 500 nm. Furthermore, the ability to collect the fibers in static or rotating collector modes allows us to generate disordered non-woven mats or single aligned fibers (Figure 2). Fluorescence microscopy, scanning electron microscopy (SEM) and atomic force microscopy (AFM) are the main techniques our group utilizes in the characterization of the electrospun nanofibers. Figure 2 shows a fluorescence micrograph of a collection of 100-300 diameter photo-luminescent polyethylene oxide nanofibers doped with two different fluorophores. Our research aims at employing electrospinning as a simple and low-cost technique for the fabrication of functional polymeric nanostructures to be used as highly sensitive biosensors for lab-on-a-chip applications.

Conclusion and outlook With the increasing demand for understanding more complex biological phenomena as well as the need for highly sensitive diagnostic techniques, micro and nanostructured materials will play a preponderant role in the study of biomolecular interactions and the development of novel sensing devices. On one hand, the development of high throughput and low-cost techniques to create more uniform and precisely controlled biomaterial micropatterns under aqueous conditions should expand the range of biological systems that can be studied and monitored in vitro. This will not only enhance our ability to study membrane and cell behavior under controlled environmental conditions,

but should also expand the range of platforms available for diagnostics based on membrane-binding analytes and for the screening of drugs that target membrane associated proteins. On the other hand, the use of simple techniques for the fabrication of nanostructured bioactive surfaces should contribute to the development of architectures that exploit the high surface area and tailored functionality provided by electrospun nanofibers, such as tissue engineering scaffolds, separation matrices, or highly sensitive electrodes.

References 1. Moran-Mirabal, J. M.; Edel, J. B.; Meyer, G. D.; Throckmorton, D.; Singh, A. K.; Craighead, H. G. Biophysical journal 2005, 89, 296–305. 2. Wu, M.; Holowka, D.; Craighead, H. G.; Baird, B. Proceedings of the National Academy of Sciences of the United States of America 2004, 101, 13798. 3. Mossman, K.; Groves, J. Chemical Society Reviews 2007, 36, 46. 4. Fang, Y.; Frutos, A. G.; Lahiri, J. Journal of the American Chemical Society 2002, 124, 2394–2395. 5. Zhou, X.; Moran-Mirabal, J. M.; Craighead, H. G.; McEuen, P. L. Nature Nanotech 2007, 2, 185–190. 6. Bally, M.; Bailey, K.; Sugihara, K.; Grieshaber, D.; Vörös, J.; Städler, B. Small 2010, 6, 2481–2497. 7. Verbridge, S. S.; Parpia, J. M.; Reichenbach, R. B.; Bellan, L. M.; Craighead, H. G. Journal of Applied Physics 2006, 99, 124304. 8. Liu, H.; Reccius, C. H.; Craighead, H. G. Applied Physics Letters 2005, 87, 253106. 9. Liu, H.; Kameoka, J.; Czaplewski, D. A.; Craighead, H. G. Nano Letters 2004, 4, 671–675. 10. Li, D.; Herricks, T.; Xia, Y. Applied Physics Letters 2003, 83, 4586. 11. Moran-Mirabal, J. M.; Slinker, J. D.; DeFranco, J. A.; Verbridge, S. S.; Ilic, R.; FloresTorres, S.; Abruña, H.; Malliaras, G. G.; Craighead, H. G. Nano Lett 2007, 7, 458–463.

Yujie Zhu is a graduate student and Dr. Jose M. Moran-Mirabal is an Assistant Professor at the Department of Chemistry and Chemical Biology at McMaster University.

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Pumps Supercritical Fluid Technologies, Inc. introduces the completely self-contained SFT-10 Liquid Carbon Dioxide Pump. The SFT-10 pump can pressurize carbon dioxide up to 10,000 psi (69 MPa) at flow rates from 0.01 to 24.0 mL/min. These characteristics make the SFT-10 an ideal pump for use in supercritical fluid extraction as well as a variety of other high pressure applications, including supercritical fluid reaction chemistry and chromatography. The SFT-10 utilizes reliable, dual sapphire syringe pump technology to achieve high pressures rapidly while its Peltier chiller has superior cooling capability. It maintains the temperature at the pump heads low enough to ensure the carbon dioxide remains liquid. The SFT-10 may be acquired as a stand alone CO2 pump or used as part of Supercritical Fluid Technologies’ SFT-100 / SFT-100XW Supercritical Fluid Extractors.

web: www.supercriticalfluids.com.

Titrator JM Science’s AQUACOUNTER® Karl Fischer Volumetric Titrator (AQV-2200S) is rugged, reliable, long lasting and eco-friendly with small volume titration cells requiring only 20mL of titration solvent for accurate measurements. Less reagent volume reduces waste and makes it easy to replace fresh solvent for the next series of measurements. This unit comes with accessories kit and is ideal for users who have many kinds of samples to be analyzed or exchange KF reagents frequently or after each measurement. A two-channel option means easy plug-and-play. End-users can add various peripherals, such as a second channel, and the system recognizes the new channels and begins working with them immediately. Intuitive control panel display shows current status, data and function keys displayed on a large, colourful screen. Result data with curves can be viewed on your PC internet browsing program without optional software. Save and load parameters and results via USB flash memory. A built-in thermal printer is easy to load and has highresolution printouts.

web: www.jmscience.com

July 2012 Laboratory Focus www.bioscienceworld.ca

New Products Columns Phenomenex Inc introduces Yarra, a new family of aqueous size exclusion chromatography (SEC) columns for biomolecule analysis. Yarra columns are offered in three phases with 3-micron particles, and are ideal for the separation of small to large proteins and peptides, as well as biological therapeutics and biosimilars. Its proprietary hydrophilic surface chemistry ensures high resolution and minimal absorption of proteins for accurate quantitation. They are also highly reproducible, column-to-column and batchto-batch. The lifetime of Yarra columns can be further extended with the SecurityGuard column protection system. Phenomenex offers complete online application and method development and optimization support for the new product.

Web: www.phenomenex.com

Chambers Sheldon Manufacturing, Inc. introduces the SHEL LAB Bactrox Chamber, the latest addition to its line of anaerobic chambers. The Bactrox is ideal for stem cell research, mammalian cancer research and clinical and research microbiology. It offers precise oxygen and carbon dioxide control from one to 20 per cent. Removal of cultures from the incubator does not require exposure of cells to undesirable oxygen or carbon dioxide levels. The unit’s advanced atmospheric controller allows for the use of a highly accurate and long-lasting zirconium dioxide oxygen sensor, with independent oxygen and carbon dioxide control and logging. The Bactrox also has a standalone 300 plate incubator so users can comfortably work without gloves in ambient room conditions. To minimize set-up time, the Bactrox includes an extra- large vacuum-less sample pass box that takes only 60 seconds to purge. The new design provides a vacuum-less sleeve entry into the chamber. Other features of this unit include temperature control and logging, superior condensation control, and ultra bright LED examination lights inside the chamber.

Web: www.shellab.com

High Throughput Developed by Union Biometrica, Inc., the Copas Plus is suitable for highthroughput analysis and sorting of human induced pluripotent stem cell (hiPS) clusters using large particle flow cytometry. Union Biometrica’s large particle flow cytometers allow the analysis and sorting of intact hiPS cell clusters from a complex mixture based on size, optical density and fluorescent parameters. This process is gentle and does not influence the morphology or viability compared to manually sorted cell clusters.

Web: www.unionbio.com

Probes Omega introduces its new pH and ORP differential probes that stay in service and provide accurate measurements under conditions that often render conventional pH probes inoperable. These probes feature integral 2-wire four to 20 mA transmitter, a built-in pre-amp that supports up to 914mm (3000’) sensor-toanalyzer distance, 4.6m (15’) standard cable length and automatic temperature compensation on pH versions. Applications include: process control, industrial and municipal water treatment, food and beverage, chemical processing, and mining and power generation.

Web: www.omega.ca

Filters Sartorius introduces its new generation of sterilizing-grade filters with the launch of the Sartopore® Platinum.With the goal of offering better filtration performance and lower filtration costs, the membrane of the Sartopore® Platinum filter cartridges has been pleated using SSB’s newly developed, proprietary TwinPleat® process. The alternating long and short pleats of the membrane increase the filter area of a 10 inch cartridge by more than 60 per cent. At the same time, this geometry ensures that liquids flow through the entire filter area so that the filtration capacity of the cartridge is used to the fullest extent. As a result, Sartopore® Platinum filter cartridges substantially boost filtration performance and lower filtration costs. The new sterilizing-grade filter cartridges are available in a choice of different sizes and constructions ranging from lab to production scale. In all sizes, identical materials of construction with consistent performance characteristics are used.

Web: www.sartorius.com

Automation Solentim introduces its new Cell Metric™ CLD bench top system for automated cell line development (CLD). Offering high resolution through cell imaging and analysis, the system offers an alternative approach to manually checking cell line monoclonality, and tracking of clones derived from single cells. The system also benefits from an automatic focus, ensuring that images are clear, consistent and informative. The Cell Metric CLD’s integrated, temperature-controlled microplate stacker allows for the automated imaging of a batch of up to 10 plates at a time, without consuming bench space. This increases throughput and reduces the need for user intervention during cell line development. With a throughput of up to 100 plates a day, the new stacker enables a substantial number of wells to be imaged and documented.

Web: www.solentim.com

Company & Advertiser Index COMPANY

Page Website

BIOTECanada...................................... 5..................................www.biotech.ca BTX.................................................. 15............................www.btxonline.com Caledon Labs.................................... 5......................www.caledonlabs.com Canada Foundation for Innovation......... 4..............................www.innovation.ca Canadian Society for.................................................................................... Chemical Technology......................... 4................ www.cehminst.ca/profdev Canadian Society for...................................................................................... Medical Laboratory Science................. 3.................................. www.csmls.org Cangene Corporation.......................... 5..............................www.cangene.com Children’s Miracle Network............ 13... www.childrensmiraclenetwork.ca Encycle Therapeutics........................... 2....................www.marsinnovation.com Eppendorf........................................ 20....................... www.eppendorf.com MaRS Innovation................................ 2....................www.marsinnovation.com Miele Professional............................ 9............... www.mieleprofessional.ca Omega.............................................. 16..................................www.omega.ca Ontario Brain Institute........................ 4.........................www.braininstitute.ca Pfizer Canada..................................... 4.....................................www.pfizer.ca Phenomenex..................................... 16.......................www.phenomenex.com Sanofi Pasteur Ltd.............................1,3....................... www.sanofipasteur.ca Sheldon Manufacturing Inc................. 16...............................www.shellab.com Supercritical Fluid Technologies........... 15............... www.supercriticalfluids.com VWR................................................. 2.................................. www.vwr.com Wyvern Scientific............................ 11....................... www.wyvernsci.com

www.bioscienceworld.ca Laboratory Focus

JULY July 9-11

July 2012

Calendar

AUGUST August 4-8

International Conference on 2012 APS Annual Meeting Cultivating Natural Bioactives Venue: Providence, RI for Health and Disease Tel: 651-454-7250 Venue: London, ON Fax: 651-454-0766 Tel: 519-263-5050 Email: aps@scisoc.org RC_lab_new:Layout 1 1/19/2012 9:27 AM Page 1 Email: info@ Web: www.apsnet.org naturalbioactivesconference.com August 5-9 Web: www. naturalbioactivesconference.com Protein Society 26th Annual Symposium July 11-12 Venue: San Diego, CA Life Science Innovation Tel: 301-634-7277 Northwest 2012 Fax: 301-634-7271 Venue: Seattle, Washington Tel: (206) 456-9567 Fax: (206) 456-9561 Email: wbba@washbio.org Web: http:// www.washbio.org/displaycommon. cfm?an=1&subarticlenbr=145

July 15-19 XXIV International Congress of The Transplantation Society Venue: Berlin, Germany Tel: 514-874-1717 Fax: 514-874-1716 Email: info@tts.org Web: www.tts.org

Email: cyablonski@ proteinsociety.org Web: www.proteinsociety.org

August 5-9 Symposium of The Protein Society Venue: San Diego, CA Tel: 301-634-7075 Fax: 301-634-7008 Email: leftwich@faseb.org Web: www.faseb.org

August 7-9 ICNFA 2012: 3rd International Conference on Nanotechnology: Fundamentals and Applications Venue: Montreal, QC Tel: 613-6953040 Fax: 613-6953040 Email: administrator@ international-aset.com Web: www.international-aset.com/ index.html

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July 18-20 Stem Cell & Regenerative Medicine: Evaluating & Delivering Stem-Cell Based Therapies from Research to Business Venue: San Francisco Tel: 312-780-0700 Ext: 224 Email: kkozub@acius.net Web: www.wplgroup.com/aci/ conferences/stem-cell-andregenerative-medicine.asp

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January 2012 Laboratory Focus

Career Spotlight Bio-economy Career Profile

Genome Canada launches competition in bioinformatics and computational biology Genome Canada and the Canadian Institutes of Health Research (CIHR) have launched a new competition they hope will lead to new technologies that will help researchers better understand the biology of living things. The idea behind the competition says Genome Canada is to create the next generation of tools needed to deal with the massive and ongoing influx of data from ‘omics research’, in particular large-scale sequencing projects. The focus of the competition will be on bioinformatics and computational biology. Specifically, bioinformatics expands the use of genomics data through the research, development or application of computational tools and approaches. It enables better ways to acquire, store, organize, archive, analyze and visualize data. Computational biology helps make sense of genomics data through computational analysis, modelling, and prediction. Competition applications are expected to propose new approaches to data analysis and data interpretation in the area of genomics, including development of software tools and algorithms. Of particular interest will be proposals addressing problems associated with current data handling and analysis and proposals addressing challenges arising from handling and analysis of data emerging from new technologies. Pierre Meulien, president and CEO, Genome Canada explains, “The genomics research community has an urgent need for efficient computational tools to collect and analyze data. Genome Canada has made it a priority to invest in this area so that key economic sectors from forestry to fisheries, agriculture to environment, energy to mining and human health have the ability to reap the full value of genomics research.” The competition is backed by $5 million in funding from Genome Canada and $1.25 million from CIHR. Of this, $4 million will support large-scale projects by multi-disciplinary teams to develop robust, user-friendly tools needed by the genomics research community. As well, $2.25 million will support small-scale projects by individuals or groups to propose innovative ideas with the potential for significant impact. CIHR’s funding preference will be in support of the smaller scale projects. The projects, in collaboration with Canada’s six regional Genome Centres, are expected to secure an additional $4 million in co-funding for the large-scale applied projects.

Compiled by BioTalent Canada Position: Executive Director of Research Name: Dr. Grant Pierce Company: St. Boniface Research Centre Salary Range: $100,000 and up per year

What I do:

My job is to coordinate research activities for the St. Boniface Research Centre and direct research activities for my own laboratory. My day-to-day activities include dealing with issues directly related to the operation of the Research Centre. I review and write grant applications, review data from the laboratory, and advise students and technicians on their research. I prepare and write articles for scientific and medical journals. I am the Chair of the Scientific Review Committee for the Heart and Stroke Foundation of Canada, which adds another interesting element. Travel is about 25 per cent of my time and becoming ever more important. The majority of travel is in Canada and the U.S., and I have the opportunity to travel to more exotic locations, like Turkey. I present keynote speeches for conferences, announcing the research in which the St. Boniface Research Centre is involved.

What education and skills do candidates need for this position?

A Ph.D. or an MD degree is required with managerial experience. Experience in clinical trial and scientific research is also required. Experiences nationally and internationally are assets in this position. An Executive Director of Research needs to be a visionary and look at the big picture. Essential skills include the ability to manage projects, people, and finances. People in this role should have organizational skills and patience, and a strong personality to act as a leader of a team. You must be part of a larger team, being a team player while communicating direction to the whole. One needs a great amount of energy to take a position such as this, as the hours can be long.

What are the best parts of your job?

The best thing about the job is the opportunity to influence more lives than any other occupation. Research into diseases such as cancer could influence millions of people and generations to come. It is an incredibly exciting field, and there is also the opportunity for great flexibility and to achieve great wealth for you and your research centre by creating intellectual property through research.

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