Journal of Laboratory Automation

We describe a novel nanostructured target plate for laser desorption/ionization (LDI) mass spectrometry, NALDI (Bruker Daltonics, Billerica, MA) target plates. The active surface comprises several layers of inorganic materials that are structured at the nanoscale and then further coated with a hydrophobic organic layer that facilitates sample deposition and LDI performance. These targets have been designed to analyze low mass (below 1500 Da), relatively polar, organic molecules and they have been shown to be up to 10 times more sensitive at detecting analytes in this range than conventional Matrix-assisted laser desorption/ionization—Time of Flight (MALDI)-TOF analysis. The targets can be used on standard LDI-TOF mass spectrometers and have been designed to fit the Bruker Flex series of mass spectrometers. This study demonstrates the utility of these targets for analyzing pharmaceutical compounds and demonstrates their superiority over conventional MALDI both in terms of performance and ease of use. Finally, we also demonstrate that these targets can be used in a unique sample preparation and analysis mode by allowing the capture and analysis of analytes from complex biological solutions.

GRAVI, presented here in its automated version, is a new bench-top sized immunoassay platform combining the advantages of microfluidics with those of simplified robotics. Characterized by dramatically reduced time to result (< 10 min) and significantly decreased sample/reagent consumption, the cost-efficient biosensor instrumentation allows performing multimenu analysis with minimal laboratory infrastructure.

Coupled to a robotic liquid handler, the system dispenses samples and reagents from conventional plates or tubes into microchannels of a microchip (GRAVI-Chip), in which assays are processed and results readout. As in conventional 96-well microtiter plates, the microchannels have a standard spacing of 9 mm to facilitate automation.

With solely gravity and capillary force-driven fluidics within the microchannels, liquids are free to flow while magnetic beads, functionalized with the antibody of choice, are trapped nearby incorporated electrodes by virtue of a magnet array. Following assay performance and electrochemical signal detection in the parallel microchannels, chips are regenerated by magnet release and rinsing of beads out from the microchannels.

Applicability of the presented immunoassay platform, delivering 100 results per hour, is exemplified here with results from the validation of an immunoglobulin assay for antibody quantification in mammalian cell cultures. Adapted to run on the GRAVI platform, this competitive assay covers a dynamic range of two orders of magnitude.

The selective desorption/ionization of analytes using nanomaterials is investigated using metallic nanoparticles. By replacing the sodium citrate capping of gold nanoparticles with self-assembled monolayers, we are able to both enhance analyte ionization and selectively capture analytes. Capping gold nanoparticles with a monolayer of 4-mercaptobenzoic acid enhances analyte ionization while greatly decreasing chemical noise resulting from alkali adducted species. Selective capture and sequential desorption/ionization of the peptide bradykinin (1–7) from a two peptide mixture is achieved using β-cyclodextrin capped gold nanoparticles. Finally, by switching from gold to silver nanoparticles, we are able to ionize both folic acid and amphotericin B. These results demonstrate that through careful control of nanoparticle surface chemistry and composition one can achieve selective analyte ionization for MS applications.

The macromolecular crystallography experiment lends itself perfectly to high-throughput technologies. The initial steps including the expression, purification, and crystallization of protein crystals, along with some of the later steps involving data processing and structure determination have all been automated to the point where some of the last remaining bottlenecks in the process have been crystal mounting, crystal screening, and data collection. At the Stanford Synchrotron Radiation Laboratory, a National User Facility that provides extremely brilliant X-ray photon beams for use in materials science, environmental science, and structural biology research, the incorporation of advanced robotics has enabled crystals to be screened in a true high-throughput fashion, thus dramatically accelerating the final steps. Up to 288 frozen crystals can be mounted by the beamline robot (the Stanford Auto-Mounting System) and screened for diffraction quality in a matter of hours without intervention. The best quality crystals can then be remounted for the collection of complete X-ray diffraction data sets. Furthermore, the entire screening and data collection experiment can be controlled from the experimenter's home laboratory by means of advanced software tools that enable network-based control of the highly automated beamlines.

As a result of a rapidly increasing demand for accurate, cost-effective, and highly flexible multiplexed assay systems, the annual market for multiplex assays is currently billions of dollars and is expected to continue to grow exponentially. Presently, there exist two broad classes of surface-based multiplex platforms—those using fixed planar surfaces (fixed arrays) and those that are particle-based (liquid arrays). We present here a description of an “Arrayable Liquid Array” platform, based on Encoded Sortable Particle (ESP) technology that combines many of the advantages of fixed and liquid arrays. ESPs have significant advantages in throughput, scalability, mixing efficiency, and flexibility over existing liquid array platforms while also incorporating the detection and manufacturing benefits of fixed arrays.

If microfluidic devices capable of rapid genetic analysis are to affect clinical diagnostics, they ultimately must be capable of carrying out more than ultra-rapid electrophoretic separations. The last half decade has seen a groundswell of activity in defining miniaturized DNA sample preparation methodologies that can be integrated with chip-based electrophoretic separations. Successfull integration of PCR-based DNA amplification and solid-phase DNA sets the stage for integrated microminiaturized analytical systems with sample in-answer out capabilities. Here we provide a brief review of the state of the art on the microfluidic integration of sample preparation processes with discussion of several systems with highly integrated capabilities, including one capable of detection of infectious agents present in complex biofluids in less than 30 min. This overview is used as a launch point to discuss the design and functionality of similar devices capable of accepting a whole blood or fine-needle aspirate sample, purifying the DNA, amplifying target sequences of the T-cell receptor-γ gene, and eletrophoretically resolving the products for detection of a signature consistent with monoclonality. We describe the details of the early experimental success in defining the individual chip-based processes required for an integrated T-cell lymphoma chip, with a vision to a device that provide sample in-answer out capabilities for diagnosing certain blood cancers in roughly 1 h.