omation mbers - Society for Laboratory Automation and Screening

TP005
Alex Berhitu
Spark Holland, Inc.
666 Plainsboro Road, Suite 1336
Plainsboro, New Jersey 08536
alex.berhitu@sparkholland.com
Denaturing Solid-Phase Extraction for Reduced Protein Interference in
Bioanalytical SPE-LC-MS
148
Co-Author(s)
Emile Koster
Peter Ringeling
Bert Ooms
Solid-Phase Extraction (SPE) coupled on-line to LC-MS can provide fully automated assays with 100%
recovery and high precision. However, many matrix compounds (e.g., proteins) are usually also trapped and can
subsequently co-elute with the drugs. Although most co-eluting proteins do not interfere with detection, they can
reduce the column lifetime or cause ionization suppression. Even though more than 95% of serum proteins are
removed by reversed-phase on-line SPE and only a part of the trapped proteins elute into the analytical system,
suppression of the MS response and reduced LC-column lifetime are sometimes reported. We have investigated
the effect of denaturing SPE conditions on the amount of eluting proteins using direct UV monitoring of SPE clean
up. To that end serum is loaded onto a HySphere C18 cartridge with acidified water and a wash step is performed
under denaturing conditions such as high zinc sulfate concentration, high and low pH and high temperature.
Results show that optimized denaturing wash conditions reduces the amount of co-eluting proteins significantly,
which means longer column life time, less pollution of MS interface and reduced risk of ionization suppression.
Compared to traditional off-line deproteinization (including centrifugation) followed by SPE, denaturing SPE is
not only fully automated but it also gives cleaner extracts. Moreover, denaturing SPE reduces the risk of coprecipitation
of drug and protein as separation of both occurs during the first SPE step.
TP006
Emily Berlin
Pall Life Sciences
600 South Wagner Road
Ann Arbor, Michigan 48103
emily_berlin@pall.com
Co-Author(s)
Michael L. Metzker, Luke Mast, Geoffrey Okwuonu and Toni Garner,
Pall Life Sciences
Kamran Usmani and Richard A. Gibbs, Baylor College of Medicine
Meeting the Increasing Challenges of Speed, Performance, and Quality While Reducing
Costs in Whole Genome Sequencing
Following the completion of the human genome sequence, the BCM-HGSC in collaboration with its respective
partners has recently released the final draft assemblies of the rat, Drosophila pseudoobscura, and honey bee
genomes. Current efforts of draft quality sequence and subsequent assembly of the sea urchin, Rhesus macaque,
and bovine genomes present new challenges in speed and performance while maintaining high quality and
reduced costs. Additionally, tracking of species-specific projects and associated BAC clones to support our
blended whole genome shot-gun strategy for Atlas assembly of whole genomes is critical for efficient project
management. Our recent innovations in automation technology in both production and instrument loading groups,
and the development of the 192-well plasmid purification platform have been instrumental in meeting these
challenges. For example, the 192-well plate format increases throughput 2-fold at reduced materials and reagent
cost while requiring the same number of research personnel and equipment (i.e., prep robots, storage freezers,
and shakers). Single plate-to-plate transfers for paired-end DNA sequence reaction set-up ensures robust tracking
efficiency of projects through our production pipeline. The current status of different whole genome projects and
the underlying production strategies to ensure the highest quality draft sequence products will be presented.