This conference follows Nucleic Acid-Based Technologies and is concurrent with DNA Forensics

Demands for miniaturization, multiplexing, and ultrasensitivity are driving genomic and proteomic research and diagnostics towards greater performance, improved labels, and other methods for detection of biomolecules. Greater speed, throughput, and accuracy are increasingly achievable, moving towards further automation and lower costs-and greater demands upon data handling and integration. Innovative techniques, including nanolabels, extended multiplexing and direct detection, help to achieve some of these goals. Challenging applications such as assays of single cells and initiatives addressing single molecule detection and manipulation help push the technologies-which also hold promise for many other applications.

Corporate Sponsor:

Sponsoring Publications:
The Analyst
Bioinform
Drug Discovery and Development
Genome Research
Genome Technology
Journal of Medical Genetics
Scanning

Scientific Advisors
Dr. Michael Natan, Nanoplex Technologies, Inc.
Dr. Monica Palcic, University of Alberta

Chairpersons
Dr. Norman Dovichi, University of Washington
Dr. Michael Natan, Nanoplex Technologies, Inc.
Dr. Shuming Nie, Georgia Institute of Technology and Emory Medical School
Dr Peter B. Simpson, Merck Sharp & Dohme Neuroscience Research Centre

Additional Speakers
Dr. Nancy Allbritton, University of California, Irvine
Dr. Mark Chee, Illumina, Inc.
Dr. Hauke Clausen-Schaumann, nanotype GmbH
Karen Cox, Eli Lilly and Company
Dr. Robert Dickson, Georgia Institute of Technology
Dr. Stephen K. Doberstein, Xencor, Inc.
Mr. Rainer Erdmann, PicoQuant GmbH
Dr. Rob Heetebrij, KREATECH Biotechnology BV
Dr. Wolfgang Kreiss, Bayer AG
Dr. Catherine Lewis, National Institute of General Medical Sciences
Dr. Wlodek Mandecki, PharmaSeq, Inc.
Dr. Jonathan O'Connell, Bristol-Myers Squibb Company
Dr. Monica Palcic, University of Alberta
Dr. Todd Peterson, Genicon Sciences Corporation
Dr. Marc Porter, Iowa State University
Dr. Salvatore J. Salamone, OraSure Technologies, Inc.
Dr. David Tabor, ACLARA BioSciences, Inc.
Dr. Dirk Ullmann, Evotec OAI
Dr. Christopher Zander, Atto-Tec

Multiplexing
Multiplexing Bioanalysis with Colloidal Metal Nanoparticles
Array Miniaturization and Highly Parallel Analysis
Solution-Phase Multiplexing of Gene Expression and Protein Analysis
Upconverting Phosphor-Based Detection
Microtransponder-Based Systems for Multiplex Analyses
Multiplexing Assays and Quantitative Biology

Nanolabels and Force Sensing
Luminescent Quantum Dots and Raman Active Nanoparticles for Multiplexed Optical Coding
Resonance Light Scattering for Sensitive Detection
Photoactivated Silver Nanodot Fluorescence
Multifunctional Metal Nanoparticles
Highly Parallel Molecular Interaction Force Assays
Atomic Force Microscopy as a Biochip Read-out Modality
Applications of Target Labeling by the ULS® Technology

Single Cells and Single Molecules
Single Cell Proteomics
Single Cell Metabolism and Enzyme Assays
Biological Systems, Quantitative Analysis, and Single Cells
NIH's Program on Single Molecule Detection and Manipulation
Single Molecule Detection in Solution

Drug Discovery and Development Screening
Practical Cell-Based Drug Discovery
Sustainable Data from Miniaturized uHTS
Imaging of Biological Activity in Living Cells
Eukaryotic Protein Display for Analyzing Small Molecule-Proteome Interactions
CLIPR and LEADseeker Imagers for 384 and 1535 LEADseeker Bead Assays

 

Wednesday, June 26

5:00- Early Registration

Additional Opportunity 7:00-9:00 Legal Roundtable Discussion DNA Match and DNA Transfer Issues

 

Thursday, June 27

7:30am Registration, Poster and Exhibit Viewing, and Light Continental Breakfast

 

Multiplexing

8:30 Chairperson's Comments
Dr. Michael Natan, Chief Executive Officer, Nanoplex Technologies, Inc.

8:40 Multiplexing Bioanalysis with Colloidal Metal Nanoparticles
Dr. Michael Natan
Au metal nanoparticles have recently become widely used components of the bioanalytical toolbox. In particular, their tunable dimensions, ease of handling, and stability are attractive, and the ability to interrogate them using a variety of physical, optical, and electrical techniques allows incorporation into a variety of detection schemes. This talk will describe two novel nanoparticles that have tremendous promise for mutiplexed biological assays: Nanobarcodes™ particles for multiplexing assays and nanoparticluate Raman-sensitive tags for multiplexed quantitation.

9:10 Array Miniaturization and Highly Parallel Analysis
Dr. Mark Chee, Vice President, Genomics, Illumina Inc.
Randomly ordered, self-assembled arrays of beads provide a novel and versatile technology platform for multiplexed analysis of complex biological samples. To form an array, beads with diameter of~3 microns are assembled into wells at the end of an optical fiber bundle. These highly miniaturized arrays have been formatted into a matrix that matches a microtiter plate, allowing many samples to be processed efficiently. The technology allows hundreds of thousands of SNP genotypes to be determined per system per day and is currently also being developed for high-throughput RNA and protein analysis.

9:40 eTag™ Reporter Assay System for Solution-Phase Multiplexing of Gene Expression and Protein Analysis
Dr. David Tabor, ACLARA BioSciences, Inc.
The eTag™ Reporter system-a universal assay platform-enables solution-phase, multiplexed assays for the measurement of both gene and protein expression in biological samples. The system enables the concurrent measurement of intracellular, membrane, and secreted proteins together with their post-transcriptional modifications. Multiplexing with eTag Reporters simplifies pharmacological profiling and deorphanization of receptor and enzyme families. The system offers rapid assay development, high-level multiplexing, direct analysis of cell lysates with no sample preparation, one-step assays with no wash steps using standard laboratory instruments, and reliable assays with high sensitivity. Key application areas include analysis of signaling pathways, surrogate markers for cell function or toxic effects, or any useful panel of mRNAs and proteins.

10:10 Poster and Exhibit Viewing, Refreshment Break

10:45 Upconverting Phosphor-Based Detection
Dr. Salvatore J. Salamone, Senior Vice President of R&D, OraSure Technologies, Inc.
Up-Converting Phosphors, or UPT, are a new class of reporter particles. These pre-ceramic materials up-convert infrared to visible light in any biologic matrix. Therefore, once labeled with probes or proteins they allow the detection of attomolar concentrations of target analyte. This presentation will review the use of these materials in a variety of formats designed to detect rare or low concentration analytes. In particular the use of these reporters for multiplexed rapid detection of nucleic acid probes for infectious diseases will be discussed.

11:15 Microtransponder-Based System for Multiplex Analyses of Nucleic Acids
Dr. Wlodek Mandecki, President and Chief Executive Officer, PharmaSeq, Inc.
Microtransponders are used in a novel DNA detection system that is capable of accurately detecting and differentiating a large number of unique DNA sequences in a single assay. Microtransponders are cube-shaped, miniature, radio-frequency transmitters, only hundreds of micrometers on each side. The presence of complementary DNA sequences in a biological specimen is determined by reacting fluorophore-labeled specimen nucleic acid with transponders, each derivatized with a different oligonucleotide probe. A scanner then detects and measures the fluorescent signal generated by the labeled specimen nucleic acid hybridized to the probe on the transponder and identifies the nucleic acid sequence involved by means of laser activation of the transponder's memory. The technology is ideal for assays in which screening for many genes, gene fragments, or mutations is necessary, a typical requirement in medicine and research.

11:45 Multiplexing Assays and Quantitative Biology
Karen Cox, Senior Microbiologist, Eli Lilly and Company
Quantitative biology in drug discovery involves the testing of hundreds and thousands of new chemical entities for their pharmacological activity. Current high-throughput testing involves biochemical and cell-based assays implemented using state-of-the-art sample processing, instrumentation, automation, and data analysis tools. Emerging areas of analytical significance in drug discovery are in multiplexing assays, miniaturization, and novel biological reagents and data informatics. This discussion will focus on the implementation of a technology for multiplexed assays and its utility in quantitative biology. We have implemented the simultaneous measurement of 5-10 secreted cytokines in 10-30 µL samples using a bead-based technology that employs fluorescent dyes as optical barcodes. Immunoassays designed using this technology (originally developed by Luminex Corporation) were used to measure various cytokines in the pg-ng/ml range in cell culture media, cell lysates, and tissue extracts. The principle of the technology, assay development, and validation issues will be discussed. Future needs and directions with respect to multiplexed assays and quantitative biology will conclude the talk.

12:15 Panel Discussion

12:45 Lunch (on your own)

 

Nanolabels and Force Sensing

2:00 Chairperson's Comments
Dr. Shuming Nie, Professor of Biomedical Engineering, Chemistry, Hematology, and Oncology, Georgia Institute of Technology; and Wallace Coulter Department of Biomedical Engineering, Emory Medical School

2:05 Luminescent Quantum Dots and Raman-Active Nanoparticles for Multiplexed Optical Coding of Genes, Proteins, and Cells
Dr. Shuming Nie
Recent advances in bioanalytical sciences and bioengineering have led to the development of DNA chips, miniaturized biosensors, and microfluidic devices (e.g., bioMEMS). These enabling technologies have substantially impacted many areas in biomedical research, such as gene expression profiling, drug discovery, and clinical diagnostics. As current research in genomics and proteomics produces more sequence data, there is a need for technologies that can rapidly screen a large number of genes, proteins, and cells. Here we report the use of multicolor semiconductor quantum dots and Raman-active metal nanoparticles for massively parallel optical encoding and high-throughput analysis of biological molecules. This technology is expected to have broad applications in genomics, proteomics, and cell screening.

2:35 Resonance Light Scattering Particles for Sensitive Detection in Bioassays
Dr. Todd Peterson, Vice President, Technology Development, Genicon Sciences Corporation
Resonance light scattering (RLS) particles are sensitive labels that have been implemented for a wide variety of analytical bioassays. Spherical gold and silver RLS particles™ of uniform dimension (between 40-120 nm diameter) generate highly intense monochromatic light when illuminated with configured white light. The colored light signal generated by a single RLS Particle is 104 to 106 times greater than the signal obtained for the most sensitive fluorescent molecule. The intensity and color of scattered light generated by individual RLS particles is stable and dependent upon particle composition and diameter according to predictive algorithms. The surface of RLS particles can be derivatized with a variety of biomolecules to affect specific binding in analytical bioassays. Sensitive RLS reagent and instrumentation systems for microarrays, immunocytology/histology, in situ hybridization, microtiter well assays, and microfluidics have been developed. Properties of RLS particles and developed bioassay applications will be described.

3:05 Photoactivated Silver Nanodot Fluorescence
Dr. Robert Dickson, Assistant Professor, School of Chemistry and Biochemistry, Georgia Institute of Technology
Utilizing simple photochemical transformations, we have produced highly fluorescent silver nanodots from otherwise nonfluorescent silver oxide nanoparticles. Readily observable on a single molecule level, these novel nanomaterials can be written with a wide array of emission colors spanning the visible spectrum. Once controlled, these materials offer promise to optically label different species with different colors and follow ensuing dynamics.

3:35 Poster and Exhibit Viewing, Refreshment Break

4:15 Highly Parallel Molecular Interaction Force Assays for Proteomics and SNP Detection
Dr. Hauke Clausen-Schaumann, Principal Scientist, nanotype GmbH
Today there is an increased need to improve the selectivity and specificity of nucleic acid and protein microarrays in order to detect slightest variations in target molecules, such as SNPs and differences protein phosphorylation. Measuring the binding and unbinding forces (tractivity) of molecular complexes, such as receptor ligand pairs or complementary nucleic acid sequences, reveals additional information on the binding complex. This extra information is orthogonal to the affinity measured in conventional binding assays, allowing for the discrimination of target molecules, which cannot be distinguished otherwise. However, typical force experiments are conducted sequentially and at the single molecule level, which makes it difficult to increase the throughput by several orders of magnitude. Here we present a new C-FIT force assay enabling us to probe large numbers of molecules (>>1010) simultaneously, using a chip format with a built-in differential force sensor that is capable of multiplexing. This force assay enables us improve the selectivity and specificity of nucleic acid and protein arrays significantly and to reduce unwanted cross-reactions.

4:45 Atomic Force Microscopy as a Biochip Read-out Modality
Dr. Marc Porter, Director, Microanalytical Instrumentation Center, and Professor of Chemistry, Iowa State University
Atomic force microscopy (AFM) has become an invaluable tool for the characterization of a wide range of interfacial architectures. In its infancy, AFM was used primarily for examining surface topographies at near atomic-scale resolution, moving more recently to mapping the composition and reactivity of surfaces with a lateral resolution on the nanometer length scale. This presentation examines the next step in the remarkable evolution of AFM: its application in the bioanalytical sciences. Results will be described that take advantage of the ability of AFM to resolve changes in surface topography in immunoassays and in monitoring enzyme activity. An investigation of utilizing nanometric-sized particles as labels will also be discussed.

5:15 Universal Linkage System ULS®: A Versatile Target Labeling Tool in Genomics and Proteomics
Robert J. Heetebrij, Manager Discovery Research, KREATECH Biotechnology BV
The ULS®, a platinum based cross linking technology, will become an important instrument within the laboratory toolbox of biomedical disciplines, since it provides a simple labeling protocol for both nucleic acids and proteins. In general, the technology has proven to be the only chemical labeling technology that equals or outperforms conventional enzymatic nucleic acid labeling procedures. In particular, in DNA microarray hybridization technology the ULS® system demonstrates one of its key advantages, the direct labeling of total RNA or its mRNA fraction. This eradicates the influence of elaborate enzymatic steps on mRNA representation, as substantiated by the data presented on cDNA expression arrays and oligonucleotide arrays. The labeling and detection of different proteins in crude mixtures by the ULS® system is another powerful feature. This is exemplified by results showing that all immunoglobulins can be labeled and detected via the ULS® in whole sera or ammonium precipitates thereof, as well as in human milk, while retaining their immuno-reactivity. The potential of this chemical labeling technology in the field of genomics and proteomics will be discussed and signifies its uniqueness as a target labeling technology.

5:45  Panel Discussion

6:15-7:30 Reception (sponsored by Cambridge Healthtech Institute)

 

Friday, June 28

7:30am Poster and Exhibit Viewing and Light Continental Breakfast

 

Single Cells and Single Molecules

8:00 Chairperson's Comments
Dr. Norman Dovichi, Endowed Professor of Analytical Chemistry, University of Washington

8:05 Time-resolved single molecule fluorescence spectroscopy in liquids and on surfaces
Mr. Rainer Erdmann, Managing Director, PicoQuant GmbH
Time-resolved single molecule fluorescence spectroscopy in liquids and on surfaces An advanced confocal laser-scanning micorscopy set-up is presented for performing time-resolved fluorescence spectroscopy measurements on single molecules in liquids and on surfaces. The system is cabable to measure emission wavelengths, polarization, lifetime, and intensity of single molecules. Various applications in chemical anaylsis and molecular biology are presented.

8:35 Single Cell Proteomics
Dr. Norman Dovichi
We have developed automated technology to generate two-dimensional electrophoresis maps from a single cancer cell. In this technology, an inverted microscope is used to characterize the cell, and micromanipulators are used to place the cell within a fused silica capillary. Simple chemistry is used to lyse the cell and fluorescently label the proteins. Automated capillary electrophoresis is used to generate two-dimensional separations, and laser-induced fluorescence is used to detect the separated proteins.

9:05 Single Cell Metabolism and Enzyme Assays
Dr. Monica Palcic, Professor, Department of Chemistry, University of Alberta
Methods are being developed to carry out biochemical analysis when sample sizes are limiting such as in the early stages of embryogenesis, after deleterious gene ablation or tumor biopsy samples. Single cell metabolism is monitored by capillary electrophoresis with laser-induced fluorescence (CE-LIF) detection after fluorescently tagged substrates are added to cell culture media, taken into cells, and biosynthetically modified. Single cell enzyme assays require miniaturization and continual sampling for CE-LIF analysis. These methods are being developed for glycosidases and glycosyltransferases involved in oligosaccharide biosynthesis; however, they are applicable to any enzyme with fluorescent substrates.

9:35 Biological Systems, Quantitative Analysis, and Single Cells
Dr. Nancy Allbritton, Associate Professor of Physiology and Biophysics, University of California, Irvine
A crucial challenge in the coming decades is to determine the functional relationships of biological molecules within the complex network of interacting proteins in a cell. In order to accurately measure the complex interrelationships of the molecules within these networks, measurements must be made in individual cells with the positive and negative feedback pathways intact. However, the technology to accurately perform such measurements is extremely limited. For this reason the Allbritton Laboratory is pursuing the development of new technologies to perform quantitative measurements of these key signal transduction molecules. These efforts include methods to measure the activation state of many enzymes simultaneously within a single cell and to quantitate the concentration of second messengers within cells.

10:05 Poster and Exhibit Viewing, Refreshment Break

10:45 NIH's Program on Single Molecule Detection and Manipulation
Dr. Catherine Lewis, Chief, Biophysics Branch, Division of Cell Biology and Biophysics, National Institute of General Medical Sciences
An overview of the recently formed NIH program on single molecule detection and manipulation will be presented. The goals of the program and the focus of NIH's interest in single molecule measurements will be discussed, including examples of currently funded research projects and opportunities for further development.

11:15 Single-Molecule Detection in Solution: A New Tool for Analytics?
Dr. Christoph Zander, Managing Director, Atto-Tec
Since the first successful fluorescence detection of a single (multiply labeled) molecule in solution by Hirschfeld 25 years ago, the research on single molecule detection and spectroscopy (SMD) in solution and on surfaces under ambient conditions has seen an explosive development. The talk aims to give an overview over the topic. In this context theoretical and practical aspects will be discussed; especially, SMD applying microcapillaries and laser diodes will be presented to demonstrate conceivable applications.

11:45 Panel Discussion

12:15 Lunch

 

Drug Discovery and Development Screening

1:30 Chairperson's Comments
Dr. Peter B. Simpson, Department of Biochemistry & Molecular Biology, Merck Sharp & Dohme Neuroscience Research Centre

1:35 Practical Cell-Based Drug Discovery
Dr. Peter B. Simpson
Rapid advances in the technologies available to imaging research are making cell imaging a major tool for drug discovery. The practical consequences will be illustrated with an example showing integrated use of high-throughput and cell-based imaging approaches to understand receptor activation. The consequences of integrating technologies of screening and cell imaging groups to improve lead and target characterization will be discussed.

2:05 Sustainable Data from Miniaturized uHTS: Efficient Approaches in Reduction of False Positives and False Negatives
Dr. Dirk Ullmann, Head, Screening Operations, Evotec OAI
The application of confocal detection resulting in multiparameter data sets improves data reliability enormously when extracting and combining the suitable result parameter. The increased data robustness enables one to perform low-threshold "sensitivity screening" runs that significantly help reduce the number of false negatives and enable accessing weak molecular interaction screening. In addition, multiparameter analysis opens the door to efficient approaches to false-positive selection by multiparameter autofluorescence filtering and detection of systematic errors. Together with readout multiplexing, confocal detection screening gives the highest precision in hit selection from primary uHTS, which will be discussed on case studies.

2:35 Imaging of Biological Activity by Living Cells
Dr. Wolfgang Kreiss, Section Head Structural Research, Bayer AG
For the imaging of the spatial distribution of biological activity we employed successfully living cells representing sensor elements of high specifity and selectivity. Ideally suited for bioactivity imaging are microorganisms, which respond to a specific stimulus by optical signals, e.g., changes of bioluminescence or fluorescence. The applications include the imaging of substance microarrays for drug discovery as well as novel chromatographic detection techniques for bioactive compounds in natural products.

3:05 Poster and Exhibit Viewing, Refreshment Break

3:30 ProCode™: Eukaryotic Protein Display for Analyzing Small Molecule-Proteome Interactions
Dr. Stephen K. Doberstein, Vice President, Chemical Biology, Xencor, Inc.
While advanced phenotyping techniques have increased the power of cell-based screening methods, the approach has ultimately been limited in utility by the difficulty of identifying the protein targets of leads identified in this way. We have overcome this bottleneck by developing affinity-based methods for profiling interactions between specific chemical entities and the entire proteome. Applications of these technologies can define therapeutic mechanism of action of new lead compounds as well as side-effect targets of existing therapeutics.

4:00 CLIPR and LEADseeker Imagers for 384 and 1535 LEADseeker Bead Assays
Dr. Jonathan O'Connell, Team Leader, Enzyme Assay Design, Bristol-Myers Squibb Company
The introduction of LEADseeker technology from Amersham Biosciences enabled a huge reduction in the read time for SPA-type assays. It became possible to read an entire 384-well plate in two minutes compared to forty minutes previously. This removed significant 384-well screening bottlenecks but also opened the door for miniaturizing these assays to 1536-well plates, increasing throughput to over 200,000 wells per day.

4:30 Panel Discussion

5:00 Close of Conference