Company Focus: SpinX Technologies

Drs. Van de Vyver and Zucchelli founded SpinX Technologies in 2003 in Geneva, Switzerland.

Overview

SpinX Technologies has developed a bench-top integrated assay assembly and detection system for assays in nanoliter volumes. Secondary screening and lead optimization subject compounds to a range of assay conditions far greater than in High Throughput Screening. Existing microplate-based technologies, well-suited for screening hundreds of thousands of compounds under identical assay conditions, fall short on providing the flexibility needed to run a large panel of subtly different assays in low volumes. Capable of running dozens of similar assays simultaneously, the SpinX Lab integrates seamlessly into today’s laboratory workflow and provides the flexibility and efficiency needed for lead optimization panels. The user simply adds reagents and compounds in the low microliter range using standard pipetting to reservoirs arrayed in a standard 384-well microplate format. The systems’ unique non-mechanical valve technology and centrifugal flow control precisely meters nanoliter volumes to potentially dozens of chambers simultaneously, enabling hundreds of different protocols that dilute, mix, and incubate. Dispensing once into a single 384-well plate can yield thousands of data points with the integrated fluorescence read-out.

Introduction

Simply miniaturizing microplate-based assays has been seen as a solution to running more samples per unit time, with the dominant format now being low-volume 384 and 1536 well microplates in High Throughput Screening. This paradigm has worked well for HTS because a large number of samples can be processed under identical conditions in parallel. Many groups have applied the same technology to secondary screening and lead optimization, with the advantages of using the same assays, liquid handling and detection instrumentation already part of their workflow. However, the parallel nature of traditional microplate-based technology is not well suited for assays probing a moderate number of compounds in a moderate number of subtly different assays.

Traditional microfluidic “lab on a chip” systems, while reducing reagent and compound consumption, are dedicated to specific applications and thus lack the flexibility needed for lead optimization panels.

The SpinX programmable technology, in contrast to traditional systems, offers an unprecedented degree of flexibility, allowing researchers to design and perform panels following the internal requirements and decision-making processes of individual lead-optimization campaigns.

Chip-to-World Interface

A critical aspect to increasing productivity and reducing reagent and compound usage is the transfer mechanism from the existing microliter-scale infrastructure to the nanoliter-scale world of the microfluidic device. For maximum productivity and efficiency, this “chip to world” interface must be seamlessly integrated with common microplate-based liquid handling systems. SpinX achieves this by presenting to the microliter-scale world what appears to be a deep 384-well microplate, called the gStack™. In reality, it is a stack of microfluidic gCards™. Each gCard has 32 input wells, with dimensions identical to those of the wells in two columns of a standard 384-well microplate. Twelve gCards are stacked edge-wise to form a complete 384-well plate. The gCards can be loaded with reagents either manually or automatically using conventional pipetting devices.

Once inside the instrument, this stack is disassembled into its 12 constituent gCards which are laid out on a rotor. Each gCard contains a network of microfluidic channels, and once the stack is loaded, the liquid is driven through these channels using centrifugal force. The instrument will run unattended through all phases of assay assembly and detection, including dilution of compounds and reagents, combination of different enzymes with different compounds, mixing and incubation, and the fluorescence readout of the assay.

Programmable Microfluidics for Assay Assembly

Increasing productivity also requires the ability to rapidly configure assays to suit specific needs. Programmable microfluidics enables sub-microliter reactions wherein the reagents, incubation steps, doses and concentrations can be chosen in real-time. This is accomplished using the Virtual Laser Valve (VLV) technology in combination with centrifugal force to move fluids on chip.

The VLV technology relies on focusing a high-radiance laser beam onto a small area on the surface of a thin foil which is specially engineered to strongly absorb laser light. The foil separates two substrates containing microfluidic structures. A single laser shot, lasting no more than a few microseconds, is sufficient to perforate the foil material. The perforation of the base creates a communication channel between microfluidic components facing the foil film. This closed-to-open transition enables the functionality of a valve.

It has been experimentally verified that the VLV is compatible with all biochemical samples of interest, specifically for genomic DNA, proteins, bacteria, yeast, and mammalian cells.

The microfluidic components have different functions: multiplexers for the choice of reagents and timing of incubation steps, and dosimeters for analog metering of volumes, assay incubation and readout. A dilution factor of up to 10 logs can be achieved for any of the inputs before entering into the final assay.

The SpinX gCards contain several rows of dosimeters, separated by multiplexers. Any assay or set of assays can be built up by moving the appropriate volumes of an arbitrary number of different liquids to specific destinations where they are mixed together before using the result in the next step. As the rotor spins, centrifugal force moves the liquid from the top to the bottom, through any available path. As soon as VLV connections are created between the two sides, additional paths become available for the liquid to flow downward.

On-Line Fluorescence Detection

In addition to providing the centrifugal force necessary to move liquids, rapid rotation of the rotor enables kinetic fluorescence detection of any number of chambers on the microfluidic gCards. The fluorescence readout is based on a combination of laser excitation and single photon counting of the fluorescence emission. The laser excitation light is focused onto a volume with a diameter of approximately 10 µm, and the fluorescence emission is collected from this same volume. As the rotor spins, the optical spot sweeps across the assay chamber, continuously collecting the fluorescence emission as a train of individual photons, with different times corresponding to different positions within the assay chamber. Simple integration of the signal allows continuous real time monitoring of hundreds of chambers simultaneously.

Secondary Screening and Lead Optimization

The first product in SpinX’s portfolio addresses the need for a convenient method to perform panels of enzyme activity assays with fluorescent readout. This is a critical need given that enzymes represent the largest target class screened. Today, in-house profiling is usually limited to a relatively small panel of less than a dozen assays, largely because no existing instrument allows a wider panel to be performed in-house in a cost-effective manner. Beyond enzyme activity assays, SpinX is developing increasingly complex assays using the same microfluidic gCards.

Programmability enables researchers to adopt and adapt a generic panel for any lead-optimization campaign: number and type of assays, procedure for suppressing false positives, validation policy for hits, use of kinetic information, determination of Km values, and so forth.

Conclusion

SpinX Lab is uniquely positioned to address the need for flexible, low volume assays in secondary screening and lead optimization. Its benefits include flexible, simultaneous assembly of many subtly different assay conditions, cost-effective compound and reagent usage by scaling to nanoliter assay volumes and seamless integration with current workflows by interfacing directly to microliter-scale 384 well microplate technologies.