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Extending the capabilities of real-time biosensing with analyte gradient injections for rapid analyte titrations and enhanced biophysical characterization
Michel Perriere, Associate Director, HYBRIGENICS
Fixed concentration injections are standard in biosensing but we show that gradient injections created using well defined dispersion conditions provide new capabilities of practical benefit in many applications.
In particular we show that well ordered gradients extending over four orders in concentration can be created with little modification to an existing flow injection system.
These Taylor dispersion injections (TDi) facilitate new assay formats that enhance the information quality of binding interaction data. Taylor dispersion theory is integrated with the standard two-compartment binding interaction model and both experimental and theoretical aspects of this new technique are investigated.
The application of the technique to complex binding, promiscuous binding, fragment screening and biophysical characterization is explored.
A new SPR approach to increase throughput
Michel Perriere, Associate Director, HYBRIGENICS
Achieving high analyte throughput while maintaining high quality kinetic data is often difficult. Increasing the number of sensing channels, or probes, is an effective means of overcoming this problem but these high channel, or probe count, systems are often confined to core facilities and often lack the sensitivity of lower channel count systems. Pilot experiments to identify the appropriate analyte concentration range for each analyte is usually required and is both time consuming and expensive. This requirement for pilot experiments may be considerably reduced by using a single analyte gradient injection (FaststepTM) that covers a wide analyte concentration range in a single injection. In contrast, conventional fixed concentration injections require a series of analyte dilutions to be prepared in order to cover an equivalent concentration range. More importantly, the overhead associated with preparing, loading and injecting each analyte dilution in such a series is eliminated. We show that FaststepTM , as implemented on SensíQ Pioneer, eliminates the need for sample dilutions, and the associated multiple injections per cycle, and produces high quality kinetics data that is equivalent to the conventional kinetics format. FaststepTM also reduces regeneration (if required) to a single injection step for a given analyte. Most importantly, FaststepTM greatly improves analyte throughput and reduces the human error associated with preparing and loading multiple analyte dilutions while reducing the complexity of the instrument routines and protocol writing.
High Content Neuronal Toxicity Assays Using iPSC Derived Neurons
Carlene Whitehead, European Marketing Communications Specialist, Molecular Devices
Neurotoxicity can cause temporary or permanent damage of brain or peripheral nervous system and has been found to be a major cause of neurodegenerative diseases. Therefore, there is a great interest in developing more predictive cell-based models and efficient screening tools for qualitative and quantitative assessment of the impact of chemical compounds, drug candidates and environmental agents. Human neurons derived from induced pluripotent stem cells (iPSC) such as iCell® neurons are very attractive for such studies because they exhibit functionality and behaviour of mature neurons, and are available in large quantities.
Here we present several assays using iCell® neurons and high content imaging for evaluation of impact different compounds on neurogenesis or neural toxicity. In a fixed cell assay, cells were stained with antibodies against neuro-specific markers ?-tubulin III and MAP2. The cells were fluorescently imaged with an ImageXpress® Micro XL whose large field of view and high dynamic range improves data quality and statistics. Positive or negative effects on development of neurite networks was assessed in real time using time-lapse transmitted light imaging, mitochondria potential, and viability dyes. We have demonstrated utility of assays showing effects of growth factors and number of neurotoxic compounds.
Shape-based profiling of fragment libraries
Francesca Sadler, Associate Director, BioFocus DPI Ltd
Fragment screening is a widely used approach for identification of new chemical series in drug
discovery projects. It provides an effective means of chemical space coverage and allows the
development of compounds that maintain good ligand efficiency.
The success of any screening campaign depends on the quality of the library which is being
screened. Fragment library collections are often generated based on physicochemical criteria,
such as the Rule of 3.1 Some consideration of chemotype div
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