LabAutomation 2006 - SLAS

MP01
Anthony Aglione
Hoffmann-La Roche
Nutley, New Jersey
anthony.aglione@roche.com
Where Laboratory Technologies Emerge and Merge
Co-Author(s)
Theresa Truitt
Ralph J. Garippa
A Streamlined Practical Workflow for Conducting High-Throughput Dose Response
and Selectivity Analysis Using High Content Screening Technologies
High content screening (HCS) translocation assays have become useful in the advancement of orphan G-protein coupled receptor (GPCR)
drug discovery programs. We utilized several integrated and workstation-based systems for an oGPCR HCS/HTS investigation to determine
receptor selectivity and EC50 values for ~400 validated hit small molecule compounds. Stable clonal cell lines of the drug- targeted oGPCR
and additional oGPCR’s used for selectivity determination were developed using Transfluor technology. The Cellomics Arrayscan-II
HCS platform was used to quantify receptor activation following a one hour incubation of cells with serially-diluted compound Our hit
criteria was established based upon the fold increase in baseline object number. Each single 384-well plate investigated a separate
oGPCR and was comprised of forty-eight baseline reference wells, sixteen maximal response reference wells (LITe assay control, ligand
independent translocation), and 10-pt dose response curves of sixteen compounds in duplicate. The process workflow utilized a Guava
technologies PCA system, a Matrix WellMate, Matrix CyBi-Well automated pipettor, Tomtec Quadra-3, and a stackable Zymark
Twister-I. Addressed topics will include the successful maintenance of stable clonal expression, cell harvesting and plating, compound
plate preparation (serial dilutions), compound addition to cell plate, and post-compound fixation, staining, and analysis of results.
MP02
Poster Abstracts
Nitin Agrawal
Texas A&M University
College Station, Texas
nitin.agrawal@chemail.tamu.edu
Co-Author
Victor M. Ugaz, Texas A&M University
A Battery Powered Compact Thermocycler for Rapid PCR
The ability to amplify DNA using the polymerase chain reaction (PCR) continues to be an indispensable tool in genomic analysis
applications including pathogen and infectious disease detection, forensics and population-scale polymorphism and mutation analysis.
We have previously demonstrated the capability to perform rapid DNA amplification in buoyancy driven closed loop reactors. Here we
demonstrate a new and advanced closed loop thermocycler (CLTC) operated by a low power miniature temperature controller. The greatly
simplified design of this system incorporates three aluminum blocks maintained at each of the three PCR temperature zones. All three
blocks are interconnected using threaded screws arranged such that when only one block is heated to 95°C (denature), the other blocks
are passively maintained at 72°C (extension) and 60°C (anneal) by heat conduction through the connecting screws. Because only one
block needs to be heated using a single heater, power consumption is minimal and the whole setup (including temperature controller) can
be operated on two ‘AA’ size batteries. Temperatures in all zones can be easily and independently adjusted by using screws of appropriate
thermal conductivities. We have demonstrated successful amplification of a 1.3kb amplicon of Lambda-DNA incorporating only 8 micro
liter reaction volume and reaction times under 30 minutes were achievable. The simplified design of this device also makes it ideally suited
for real time PCR and for DNA sequencing applications.
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