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High Throughput Screening CoreIntroduction to High Throughput Screening CoreThe Screening Core is a centralized, staffed facility capable of medium-to-high-throughput screening in support of biological researchers. The infrastructure within the Screening Core enables multiple assay formats and readouts, designed to produce the most relevant experimental outcomes for a given project. The Screening Core enables screening techniques most relevant to the BU research community, and features not only standard, target-based screening techniques (such as enzymatic assays, radiometric or spectrometric binding assays, and receptor-coupled assays), but also a functional screening platform (fxHTS), a whole cell-based assay that measures cellular endpoints, such as secreted proteins, fat content, or metabolic markers such as O2 consumption, CO2 generation, pH variation, etc. The Screening Core will provide equipment, expertise, and training to researchers within the University who wish to utilize the Core. A key aspect of the Core will be training; all screens will be designed, optimized, and executed by the visiting researcher in collaboration with the Screening Core staff. Funding Opportunities:
Education:Learn about Drug Discovery in the spring course CH744 Contemporary Drug Discovery HTS Core Seminar:Title: Non-Invasive Assessment of Unlabelled Primary Human Cell Cultures using the MIAS-2 HCA reader Using fluorescent techniques, High Content Analysis (HCA) has been developed to automate the quantification of complex biological phenomena inside cells, like protein activity in signaling pathways. With the aid of fluorescent protein constructs, HCA based compound screens have been developed and used. However, the need for generic assays to read additional endpoints for the drug development decision chain remains eminent. Preferentially such assays can be combined in the functional screening protocol. We have applied automated brightfield imaging to unlabeled, living cultures of skin-derived primary epidermal keratinocytes using the MIAS-2 microscopy reader. This rapid non-invasive cell quantification can be used to quantify the modulating effect of drug-like compounds on cell growth and cell death. The generic assay format allows for assessment of stimulating or ADME-TOX in combination with a wide variety of cell based screens and model organism assays. The non-invasive methodology allows to efficiently add these additional endpoints into existing screening protocols. Mission of the BU High Throughput Screening Core:
Screening Core Project flow
The typical project workflow, as shown in the figure, consists of four steps: Project design. The biological investigator (BI) brings a project idea to the Screening Core staff and discusses their screening needs, hypotheses and desired outcomes. Together, the Core and the BI develop a project plan and sets project plans, goals and milestones. Screening design. The Screening Core scientific team assists the BI’s research team in designing and validating high throughput screens, with the bulk of the assay development and validation work performed by a researcher from the BI’s research team. This will be validated by performing a small subset screen (~5k compound). Data is initially generated using a positive and negative control, generating 100-400 assay points for each. The mean and standard deviations for the two controls are calculated, and a z factor (a measure of assay robustness) is generated. Based on this, the Screening Core can assess if the assay performance is suitable for screening, and the hit criteria are established. Screening execution. The screen is executed by the Screening Technician at single compound concentration (~1uM) per the design plans, screening against the compound screening set (~25k compounds) obtained from the BU Compound Repository. Screening data for each compound is deposited in a database (IDBS, ActivityBase), which is mined for compounds passing the hit criteria. Those that meet these criteria will be retested at a single concentration in order to confirm the hit. In the function-based screen, cell health is assayed at the end of the assay using Promega's CellTiter-Blue reagent. Compounds that impair cell viability are eliminated based on the fluorescence of alamar blue, a measure of cell viability that can be assessed following determination of function and on the same cells. Confirmed screening hits are assayed at varying concentrations of compound to establish the ED50 (dose that generates half of the max effect). Screening hits that produce a sigmoidal dose response curve are re-assayed to confirm the ED50. Hit triage. Working with the cheminformaticist, the director of the MCCSF-BU assesses the hit matter obtained in the high-throughput screen on the basis of several criteria, including potency, chemical tractability and attractiveness, physical properties, and prior art in the area. Active hit compounds are re-procured by purchase or synthesis and the purity and identity checked (LCMS, NMR). Those that are confirmed pure and active are designated as “confirmed hits”, and rank ordered on the basis of efficacy and potency in the screening assay. These compounds will be provided to the BI and flagged for potential collaborative follow-up within the BU-CMD. ContactsDirector of Screening Technical Director Location670 Albany St., 2nd Floor Director Medicinal Chemistry and Core Synthesis Facility (MCCSF) Location590 Commonwealth Avenue, Instruments/ServicesUse of the devices/equipment in this core is by request and organized by an online scheduling program. To gain log-in credentials, please follow this link to complete the online application form.Small Molecule ScreenAssays will be designed and optimized in the lab of the biological investigator. The responsibility for the screen will transfer to the screening facility once the screen has been validated, automated and miniaturized. The Biological investigator is required to participate throughout the screening campaign. Rate: BU investigators $1 per compound (5000 compound minimum). This rate includes help and advice from the screening facility staff throughout assay development and during the screening campaign, use of the equipment during the screening campaign, use of the desired compound library at a single compound concentration, and hit triage. Assay reagents and supplies are the responsibility of the biological investigator. Compound Library
The compound library is only available to investigators who run their HTS campaign at the BU HTS facility. Plate Reader - Tecan, SpectraFluor PlusSchedule this device (Log-in credential required) The SPECTRAFLUOR Plus is a fully automatic, computer controlled Fluorescence, Luminescence and Absorbance instrument for measuring samples in a microplate. The filters are arranged in filter slides; four excitation filters and four emission filters respectively can be installed in the slides. Spectral range 360-700nm for fluorescence. 230-750nm absorbance spectral range. Sensitivity =1.5 picogram fluorescein on each well or =4 fmol well. The emission signal is measured by a highly sensitive PMT. Thermal incubation at +5: C to + 40: C. Dual optical channels allow measurements from above or below the plate by mouse-click. Light to and from the samples is focused by a lens and high quality filters are easily accessible from the front of the instrument. Rate: BU: Self - $10/ hr, Assisted - $30/ hr Plate washer - Bio-Tek Instruments, ELx50 Auto Strip washerSchedule this device (Log-in credential required) The ELx50 Auto Strip Washer is a self contained and programmable instrument, allowing for full control of precise fluidic delivery from the gentle dripping of a simple squeeze bottle to the full force of pressure delivery systems. Rate: BU: Self - $10/ hr, Assisted - $30/ hr Letter of Support |
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