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The Laboratory Robotics
Interest Group Mid Atlantic Chapter |
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The Laboratory Robotics Interest Group
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ABgene North America |
IDBS IGEN International Intermountain Scientific ISLAR Isthmus Kendro Laboratory Products Labcon Labman Automation LabVantage Solutions LEAP Technologies Luminex Marsh Biomedical Products MatriCal Matrix MDL Information Systems Mettler-Toledo Bohdan Millipore MJ Research Modern Drug Discovery Molecular Bio-Products Molecular Devices MWG Biotech Nalge Nunc International Nichiryo America NuGenesis Oyster Bay Pump Works Packard BioScience PerkinElmer Life Science Personal Chemistry Pierce Endogen pION |
Process Analysis & Automation Promega Pyrosequencing Qiagen Instruments Remp Robbins Scientific RTS Thurnall Scientific Software Society for Biomolecular Screening SPEX CertiPrep Spike International Spotfire ST Robotics International Tecan Technology Workshop TekCel The Automation Partnership Thermo Orion Thermo Savant THK America Titertek Instruments Tomtec Universal Imaging Upstate Biotechnology USA Scientific Velocity11 VelQuest Whatman Xyntek Zymark |
3:30 PM, Piscataway Room
Accurate, Automated SNP Determination Using the READIT?System
Dr. J. Shultz
Promega Corp. Madison WI 53711
Accurate, automated SNP determination will be needed to take advantage of the information that will arise as a result of the Human Genome Initiative. In this presentation, a system will be described that utilizes two enzymatic reactions to generate a light signal through the action of Luciferase to indicate if a particular DNA sequence is present in a sample. The first enzymatic reaction depolymerizes a DNA primer to generate dNTPs if the primer hybridizes to the target sequence without mismatched bases. The second reaction transfers the terminal phosphate of the released dNTPs to ADP to form ATP. The amount of ATP made is then measured using a Luciferase-based reaction. Comparison of the signal strengths for reactions performed to detect both alleles of a known SNP allows the user to determine the particular genotype of the sample. Data will be presented on different studies using thousands of samples demonstrating that the system is very accurate and reliable. This presentation will also describe the various robotic systems that have been used to perform READIT genotype determinations that allow the method to be performed to satisfy a variety of throughput needs.
4:00 PM, Piscataway Room
ICASTTM: Monitoring Protein/Protein Interactions as a Functional,
Cell-based Assay for GPCRs and other Membrane Receptors
Melissa Gee
Applied Biosystems
Receptor-mediated cell signaling is dependent upon specific protein/protein interactions that allow a cell to respond appropriately to an outside stimulus. We have developed a method, ICAST (Intercistronic Complementation Analysis Screening Technology), that allows one to monitor these protein interactions in a cell-based format thus providing a functional read-out for receptor activation. In this system, the interacting protein partners are expressed as fusion proteins to two different complementing b-galactosidase mutants. Enzyme complementation is contingent upon the interaction of the target protein pair such that b-galactosidase activity can be monitored to indicate protein/protein interactions. The fusion proteins are expressed at relatively low levels in mammalian cells allowing for assay conditions under near physiological conditions. By using this system to probe interactions of GPCRs with down stream signaling partner proteins, we have developed a functional assay that allows for both agonist and antagonist screening. We will present HTS data generated for the b2-Adrenergic Receptor using this system, as well as functional data for the chemokine receptor CXCR2. We have also used ICAST to monitor EGF receptor ligand-dependent dimerization in a high-throughput format, and to monitor down stream signaling events. This system provides a cell-based, functional assay that can be used for target validation and high-throughput screens for lead discovery and optimization.
4:30 PM, Piscataway Room
The Science of Discovery meets the Business of Discovery with LabVantage
Sapphire Informatics Edition
Dr. Jim Bower, Informatics Group Leader
LabVantage Solutions
Dr. James Bower has a Ph.D in Analytical Chemistry from the University of Florida. He has extensive experience in designing and applying decision support information technologies and applications to facilitate engineering and compliance management.
LabVantage Sapphire Informatics Edition, has been developed to meet the needs of high throughput laboratories. By integrating different information management systems, software packages, instruments, external enterprise databases and robotic control systems within a single auditable database, LabVantage has created end-to-end drug discovery solution. Dr. Bower's presentation will include a high-level review of the Informatics Edition's functionality including:
* Daughter plate production
* Reformatting, re-racking & cherry picking for 96/384/1536 plate
types
* Membrane/filter media
* Assay/compound plate mapping
* Automated robot scheduling
* Seamless integration with leading robotic solutions
* Graphical drag and drop workflow process capability
* Visualization of laboratory workload bottlenecks for resource planning
6:00 PM, Piscataway Room
Zymark New Product Offerings and Innovative Solutions
Jeff Donahue
Zymark Corporation
There have been numerous advances in instrumentation and compete systems that have facilitated in the acceleration of microplate manipulation and assay throughput for the purpose of streamlining the drug discovery process. Greater scrutiny has placed on the scientist for the procurement of instruments and systems in order to collapse timelines in efforts to get to the market faster, thereby saving money.
Zymark, a leader in innovative automated solutions for drug discovery is introducing the concept of the minisystem. The foundation of the minisystem is joining of Zymark's two latest products, the SciClone ALH and the Twister II. These two instruments are able to work seamlessly together through Zymark's' revolutionary open architecture software CLARA?/b> or PAM?/b>. This talk will focus the features and attributes of the recently launched SciClone ALH, Twister II and the ability to scale solutions using CLARA and/or PAM open architecture software.
6:00 PM, Edison Room
Luciferase Technology for High Throughput Screening Applications
Erika Hawkins, MSc Promega Corporation, Madison, WI
Luciferase technology is widely used to provide simple, rapid, and sensitive quantitation of cellular and other biological activities. Recent advancements in luciferase technology can be viewed through the interrelationship of two distinct areas: the genes encoding the luciferases, and the corresponding assay reagents for quantitating bioluminescence. Novel synthetic reporter genes encoding firefly, Renilla, and click beetle luciferases have been created for maximizing sensitivity and reducing the risk of anomalous expression. These new genes are essentially devoid of transcription factor binding sites, or other sequence motifs, that could interfere with reporter function. For high throughput quantitation of gene expression, homogeneous reagents have been developed to maximize assay precision and sensitivity in high-density multi-well plates. The interrelationship between gene and reagent is exemplified with Renilla luciferase. Luciferase expression from the new synthetic gene is typically increased more than 10-fold over the wild-type gene. Sensitivity may be further increased over 100-fold by proprietary methods for minimizing background in the assay reagents. In conventional reagents, the quantitation of Renilla luciferase can be compromised by background due to autoluminescence from the reagents. New assay reagents are being developed for Renilla luciferase that virtually eliminate this background. Included in these are reagents for dual reporter applications, where the firefly and Renilla luciferases are combined into a single analysis. Also under development are genes and reagents allowing multiplexed analysis of differently colored luminescence. In addition to high throughput analysis of gene expression, luciferase technology is also being applied to rapid quantitation of cell viability. The new CellTiter-GloTM Reagent provides linear correlation of luminescence to cell number over a range of 50 to 50,000 cells. In a single step, the reagent lyses cells, inactivates endogenous ATPases, and generates a stable luminescence signal proportional to intracellular ATP. This new reagent is designed specifically for high throughput screening, providing faster results with greater sensitivity than conventional reagents.
6:30 PM, Piscataway Room
Managing Proprietary Compounds to Accelerate Drug Discovery
Richard Lane
EMAX Solutions
To speed research for the next blockbuster drug, discovery organizations are investing in advanced production and testing technologies such as genomics, combinatorial chemistry and high throughput screening systems. Point solutions are mass-producing up to a half-million discrete samples a day, creating a blessing in terms of throughput, but a burden in terms of materials management. Proprietary compounds and natural products must be registered, inventoried, prepared, shared, tracked, tested and retested. Managing this immense collection of discrete substances can quickly become a logistical nightmare that diminishes the odds of discovering a potentially viable drug candidate.
This case study addresses the critical bottlenecks in drug discovery programs generated in great part by high throughput technologies. We demonstrate the necessary components that will link end users into the high throughput discovery process and support the just-in-time supply, management and tracking of substances necessary to meet the compound collection management needs of research organizations.
6:30 PM, Edison Room
An overview of Coherent Synthesis(tm), a new concept for faster chemistry
development, recently launched by Personal Chemistry, Inc.
Andreas L. Hoel has a M.S. in Chemistry from the Technical University of
Denmark. He has worked with several aspects of microwave-assisted chemistry,
including investigation of new reaction types and development of new
instrumentation. Presently he is Chemistry Group Leader with Personal Chemistry,
Inc., Milford, MA.
Coherent Synthesis, based on microwave technology, has been developed to address one of the main bottlenecks in the post-genomic era; the creation of new chemical substances for drug development. By use of automated synthesizers as well as kits for reaction optimization and custom designed software, Coherent synthesis speeds up the chemistry development process and expands the range of chemical transformations plausible.
7:00 PM, Piscataway Room
MicroWell to MicronArray: Utilizing Modified Surfaces with Substrate
Formats, Geometries and Optical Properties to Achieve Optimal Performance in
High Density MicroWell?and MicronArray?Assay Development
Barbara M. Sullivan, Ph.D. is the Director of Life Science Discovery Products at Nalge Nunc International. She has over 10 years experience in developing and fine tuning assays for diagnostics and discovery as well as in developing products to be used in various assays as the Director of Research, Quality and Technical Support for Nunc, Inc. and NNI. She received her Ph.D. in Biochemistry from Michigan State University and performed postdoctoral research in the Department of Human Genetics at the University of Michigan.
An understanding of a reaction vessel s molecular surface and physical characteristics is critical to achieving sensitivity and specificity in high throughput or high content genomic and proteomic assays. The molecular and atomic compositions of various MicroWell surfaces were defined through Electron Scanning Chemical Analysis (ESCA or XPS). Contact angle and surface charge of various reaction vessels were determined. Various molecular surfaces, in conjunction with the physical properties of the substrate material (e.g., optical or thermal properties) and format (MicroWell, micronarray, VirtuWell?slide, or chip), were compared with regard to assay performance. Well geometry, round, square, columnar, shallow or a VirtuWell 2D format influenced assay performance, particularly when an active surface was utilized. Additional parameters came into effect when well volumes were miniaturized as in a 1536 well plate. The effects of the molecular surface and physical characteristics such as optics and hydrophobicity on assay sensitivity and specificity changed again as the assay was further miniaturized to a two-dimensional slide or chip format. Covalent binding surfaces proved to be extremely effective for microarray based assays, allowing one-pot assays, easy handling and effective use of hybridization conditions to eliminate non-specific background and increase sensitivity. Fluorescent and reflective properties of solid black, solid white and black or white optical bottom plates (OBP) were compared by measuring signal to noise ratios, light cross talk and sensitivity. This data will be used to demonstrate the specific utilization of surfaces and formats in molecular and cell-based assays.
7:00 PM, Edison Room
APPLICATIONS OF HIGH-THROUGHPUT PHARMACEUTICAL ANALYSIS USING TURBULENT
FLOW CHROMATOGRAPHY / TANDEM MASS SPECTROMETRY (TFC-MS/MS)
Dr. Daniel Magiera, Senior Applications Scientist
Cohesive Techologies
Dr. Daniel Magiera is a Senior Applications Scientist at Cohesive Technologies, Inc., Franklin, MA. Dr. Magiera has more than 10 years experience in sample preparation techniques used for LC-MS/MS and GC-MS applications. He received his doctorate from Northeastern University where his doctoral thesis involved sample cleanup procedures such as solid phase extraction (SPE) and satellite HPLC to purify derivatized DNA adducts for trace detection using GC-MS. Before joining Cohesive, Dr. Magiera served as Senior Staff Scientist at Muro Pharmaceuticals/Asta Medica developing solutions for pharmaceutical methods development using HPLC-MS/MS.
LC-MS/MS plays a major role in the analysis of drugs during the discovery process. The sensitivity and specificity of LC-MS/MS has made it a preferred method for quantitatively measuring drug concentrations in plasma. With an increasing demand for high sample throughput, preparation of plasma samples prior to LC-MS/MS analysis often prevents this methodology from being a high throughput technique. This presentation describes turbulent flow chromatography, which enables the direct injection of plasma samples into a LC-MS/MS system. Unique features of this technology, which eliminate the need for sample preparation procedures such as protein precipitation, liquid/liquid extraction, and solid phase extraction, are presented. The time advantage of eliminating sample preparation is compounded with analysis times as short as 60 second cycle times for high sample throughput. Examples are shown of single and multiple analyte measurements of pharmaceutical compounds from direct plasma and urine injections. Advantages of TFC methods development and validation will also be discussed.
7:30 PM, Piscataway Room
Novel applications of a biological label for target validation and lead
identification
Richard M. Eglen
DiscoveRx Corp., Fremont, CA 94538
Enzyme fragment complementation (EFC) of beta-galactosidase (beta-gal) occurs when an alpha-donor peptide (ED, enzyme donor) restores activity to inactive, alpha-acceptor mutants (EA, enzyme acceptor). ED thus acts as a biologically inert tracer, suitable for labeling ligands (large proteins and small organic molecules), either recombinantly or chemically. DiscoveRx technology , using this approach, facilitates homogeneous assay development for several targets, including enzymes (e.g. ser thr kinases, tyrosine kinases, tyrosine phosphatases), receptor ligand binding (e.g. nuclear receptors), and second messengers (e.g. cAMP, cGMP). Furthermore, novel uses of the biological label, ED, allow interrogation of target interactions, difficult to assess otherwise. These include protein : protein interactions, protein allosteric modulatory sites and site-specific inhibitor binding (e.g. PKC - staurosporine) and novel formats for proteases, nucleases and helicases assays. Collectively, beta-Gal EFC allows assays with all the attributes required for HTS, both for inhibitor identification, and high throughput detection of novel protein : protein interactants.
7:30 PM, Edison Room
A STRATEGY FOR AUTOMATED DATA MANAGEMENT AND 21 CFR 11 COMPLIANCE
Mary Ellen Goffredo
NuGenesis Technologies Corporation
Machine-readable data, such as raw, result and method files acquired from analytical instruments, and human-readable data, such as reports generated by instrument data systems and common business software applications, are key components of electronic records. However, due to the wide variety of analytical techniques currently in use, the management and ultimate safekeeping of these data is becoming more and more problematic.
This talk will describe application independent technology solutions that may be deployed for the specific purpose of automatically managing and maintaining data backup, archival, long term storage, and retrieval of electronic records in a 21CFRll compliant environment. Archived data is catalogued in a highly searchable database using context sensitive tags that allow the data to be located easily using simple search tools, thereby ensuring access to the data by all users. Specific examples of record maintenance in an application independent environment within the pharmaceutical industry will be discussed.
8:00 PM, Piscataway Room
Semi-Preparative High Throughput Separations via a Parallel HPLC
Configuration
Joan M. Stevens, Ph.D., technical support manager and CyberLab technical
specialist
Gilson, Inc.
Analysis via HPLC contains three components: sample preparation, chromatography, and data analysis. If chromatography is the rate-limiting step, chromatographic speed can be optimized to further increase throughput. The next logical step to increasing throughput would be the addition of more chromatographs to the analysis process. The challenge as more chromatographs are implemented in the system is to handle and coordinate samples, chromatographs, fractionation, fraction tracking and data from multiple HPLC systems.
We explored optimization of sample throughput via a single multiple probe/injector device configured with multiple chromatographs (4), employing fast HPLC separations, semi-preparative flow rates, independent fraction collection, fraction tracking and parallel analysis. With this approach, semi-preparative HPLC analysis of milligram quantities (20-40 mg range) was achieved in less than 7.5 minutes per set of four samples or a series of 32 semi-prep samples were chromatographed and fractionated in an hour, or 256 semi-prep samples per day. This presentation will cover key aspects of hardware strategy, system control, chromatographic performance, sample/fraction tracking and data handling.
8:30 PM, Piscataway Room
Xyntek's Drug Discovery & Laboratory Automation Group offer consulting
and engineering services to address the following issues:
1. Process and Automation Gap Analysis
Xyntek will complete the user interviews, process assessment, and hardware
inventories necessary to define both the As-Is and To-Be drug discovery system
definitions. The Gap Analysis will provide a concise statement of high level
technical options.
2. Technical Strategy and Project Definition
Based on the client's business objectives Xyntek will prepare detailed technical
and project specifications for implementing specific automation tasks including:
Robotics, Liquid Handlers, Machine Vision, Refrigerators, Bar-Code, Scales,
Printers, SCADA, Data Collection, etc.
3. Project Implementation and Support
Xyntek will provide the project management and engineering services necessary to
deliver turnkey drug discovery and laboratory automation solutions.
We will present an overview of our Compound Management and HTS Fulfillment
Software Platforms:
?WEB-Based Ordering
?Initial Compound Distribution
?Inventory Management
?Integrated Weigh Station
?Mother & Daughter Plate Production
?Order Tracking & Related Informatics
?Other HTS Fulfillment Options
?Machine Vision/ 2D Marking of 96 Well format
Implementation of an Industrial Process for High Throughput Genotyping of
Single Nucleotide Polymorphism by Single-Base Primer Extension
Felicia A. Watson, Michael S. Philips, Leslie Picoult-Newberg, Mark G. Pohl,
Denis M. Grant and Michael T. Boyce-Jacino
Orchid BioSciences, Inc. Princeton, NJ 08540
The MegaSNPatron: An Ultra High Throughput Genotyping Platform
Craig A. Gelfand, Carol Emerich, Yi Liu, Kathryn E. RAyca, Shobha, Varde, Bahar
Hoghooghi, Mark G. Pohl, Gary Schnerr, Frank Shemansky, Rolf E. Swenson, Joel T.
Tarantino, and Michael Boyce-Jacino.
Orchid BioSciences, Inc. Princeton, NJ 08540
High Throughput Education and Training Initiative at Rutgers
Gerben J. Zylstra, Professor of Biochemistry and Microbiology
Biotechnology Center for Agriculture and the Environment
Foran Hall, Cook College, Rutgers University
High-throughput chromatographic data analysis using client/server based
Chromatography software and a proprietary visual basic application module
Christian Eckhoff 1 , Xianqun Wang 1 , James Schibler 2 , A.W. Fitchett 2 , 1
Cerep Inc., Redmond, WA, USA; 2 Dionex Corporation, Sunnyvale, CA, USA
In order to determine the aqueous solubility of small organic molecules, an HPLC procedure has been developed that is capable of analyzing hundreds of injections of chemically diverse compounds daily on one instrument. Assay results consist of aqueous solubility in the range of 1-200 然, purity (relative peak area at the detection wavelength), and UV-VIS spectrum of the compound. We faced two great challenges during assay development: 1) Information about the molecular structure of compounds to be analyzed in this assay is routinely not disclosed by Cerep's customers; 2) Sequence programming and data analysis could easily become rate-limiting for productivity when several hundred injections are performed every day. Point 1) was successfully addressed by developing an ultrafast universal HPLC method (total run time: 3 min per injection) with wide-bandwidth PDA detection. The enhanced workflow of Dionex' Chromeleon software and a Visual Basic application in Excel developed at Cerep have made it possible for the assay operator to prepare, program, inject, analyze, and audit chromatographic results of up to 200 compounds or 400 injections daily. At the same time, chromatographic data are archived, results are electronically submitted for QC-approval, and client reports are prepared.
Homogenous Assays for Nuclear Hormone Receptors using Enzyme Fragment
Complementation
Peter A. Fung, Yali Shu, Riaz Rouhani, Rueyming Loor, Richard M. Eglen
DiscoveRx
Corporation, Fremont, CA
We have developed homogeneous, in vitro assays for nuclear hormone receptor ligands using ?galactosidase fragment complementation (HitHunterTM EFC Assay Format).
In this homogeneous binding assay, two inactive fragments of E. coli ?galactosidase, termed Enzyme Acceptor (EA) and Enzyme Donor (ED) were used. In solution, EA and ED rapidly complement to form active tetrameric enzyme. Progesterone or estrogen was conjugated to ED in such a manner that does not affect complementation. In the assay, inhibitors of steroid receptor binding displace the steroid - ED conjugate, allowing complementation with EA, and a consequent increase in formation of ?galactosidase. Enzyme activity is, in turn, detected using either a chemiluminescent (dioxetane-galactoside) or long-wavelength fluorescent (resorufin-galactoside) ?galactosidase substrate.
Studies with human estrogen a receptor were undertaken using recombinant material. A crude preparation of the human progesterone receptor was prepared from Sf9 cells infected with a progesterone receptor construct. Specific binding of either estrogen or progesterone - ED conjugates was saturable, specific and Scatchard analysis revealed affinities (KD) of 2 and 0.1 nM, respectively. Specific ED - estrogen binding was competitively displaced by several ligands with the following rank order: b estradiol > diethylstilbestrone > esterone > tamoxifen > 4 OH tamoxifen. Specific ED - progesterone binding was competitively displaced by several ligands with the following rank order: progesterone > 4 pregnen 20 a, 21 diol one > 17 OH progesterone > 4,6 pregnadien 6,17 dimethyl 3, 20 dione > 6 a methyl 17 hydroprogesterone.
This simple homogenous, two-step reagent addition assay, performed in 384 well plate formats, is highly applicable to automation, possessing good precision (Z' values of 0.5 - 0.7) and CV values of 4 -6%. We conclude that the HitHunter estrogen or progesterone assays provide a robust, simple assay for nuclear receptors with performance characteristics suitable for high throughput screening.
An electrochemiluminescence-based assay for cAMP
T. Shafer, J. Schmidt, J. Karaszkiewicz and E. Kenten
IGEN International, Inc.
Comparison of Microplate Sealing Tapes Using Standardized Test Protocols
Terry W. Lewis, PhD, Maurice H. Kuypers, Mialena M.
Walker Medical Specialties Department - 3M Health Care
Adhesive tapes have become an attractive option to cover microplates in bioanalytical, genomic, and pharmaceutical research. Primary performance criteria for microplate adhesive tape seals include: prevention of evaporation from the individual wells; low contamination of well contents by the tape adhesive; prevention of cross-contamination between individual wells, and clean tape removal for access to well contents. Depending on the particular research objective, other criteria for adhesive tape seals may be important. These include pierce-ability for access to individual wells without entire tape removal, good optical properties, for monitoring well contents through the tape, and temperature resistance over wide ranges to include compound storage and PCR. This poster will describe results of a wide variety of adhesive tape performance testing designed to mimic actual industry use conditions. The tests include evaporation data with water, DMSO, and aqueous mixtures of isopropanol, methanol, and acetonitrile. Microplate composition and manufacturer were also varied. Temperature conditions ranging from -70?C up to room temperature and actual PCR cycling were evaluated. Other testing results on adhesive extractables, tape optical properties, and tape application and removal will be presented.
Determination of Single Nucleotide Polymorphism using High Throughput Mass
Spectrometry-Based System
B. Darnhofer, et al.
Sequenom, Inc.
The Society for Biomolecular Screening Microplate Standards Working Group
Date: Monday June 4, 2001
Time: 1:00 pm - 3:00 pm
Place: Hilton hotel in East Brunswick, NJ, the Edison Room
Notes: The meeting will precede the June meeting of the Mid Atlantic Chapter of
the Laboratory Robotics Interest Group who have graciously provided the room.
Advanced Technology seminar hosted by Beckman Coulter, Inc.
"Smart Solutions for Drug Discovery"
Please join us for an Advanced Technology seminar hosted by Beckman Coulter, Inc. focusing on innovative approaches for automated assay development, primary and secondary screening and 'High Content Screening' with Cellomics. You will have the opportunity to meet other users and discuss their automation applications.
Accelerated Assay Development and Optimization
- Statistical Design of Experiments
- Smart Robotics
- Data Management
Primary and Secondary Screening
- Compound Screening
- Multiple Measurements of Cellular Processes
- High Content Screening
- Informatics
June 4, 2001, 11:00 am - 3:00 pm, Hilton Hotel, 3 Tower Center Blvd., East Brunswick, NJ 08816
To register for the seminar, to view agendas and to obtain abstract
information, go to
http://www.beckmancoulter.com/seminars2001
The first class Hilton Hotel is located at the crossroads of New Jersey's major arteries at Exit 9 of the NJ Turnpike, just outside New Brunswick. It's central location is just 25 minutes from Newark Int'l Airport, 45 minutes from New York City and less than an hour from Philadelphia. Easy access to all surrounding business areas and attractions makes the Hilton East Brunswick the perfect location for business, pleasure or meetings.
On-line directions may be found at:
http://www.hilton.com/en/hi/hotels/directions.jhtml?ctyhocn=EWRBHHH
Visit The Laboratory Robotics
Interest Group homepage at https://www.lab-robotics.org
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