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High Throughput And High Content Screening What Is HTS ? Biochemical target Test many compounds Compounds with desired effect on target = Hit Hit directly used as chemical probe to study target OR Hit provides structure for chemists to begin developing a drug or other product HTS Technology Robotics Miniaturization Sophisticated Assay Chemistry Sophisticated Software and Database Evotec uHTS System • Opera High Content Detector • Plate:vision detector for fluorescence (also fluorescence polarization & fluorescence resonance energy transfer), luminescence, absorbance • 2 CO2 incubators • 4 low volume pipetting systems for 384and 96-well plates (multidrops, Cy-Bi well, Cy-By drop) • Plate washer • Centrifuge • Piercer • Plate storage, barcode readers, lid handlers, transport systems plate::vision detector • • • • • CCD camara Xenon lamp light source Zeiss optics Built in algorithm to correct for spherical lens aberration 4 filter sets – Excitation filter – Emission filter – Dichroic filter Opera 4 lasers 1 xenon lamp 4 CCD cameras 96- and 384-well plates Acapella image analysis software Software Systems Genedata • Assay Analyzer • Condeseo • Integrated with Opera Acapella image analysis software Accelyrs Accord • HTS • With Pipeline Pilot Ohio Third Frontier Computing Systems HTS Output is QUALITY DATA Standard Screening Procedure 1o HTS hits Triplicate assay confirmed HTS hits 10 point dose-response assay EC50 HTS Assay Examples • Receptor-binding • • Enzyme • • • • • • • • GPCR protease, kinase, phosphatase, lipase, others Bacterial growth Cell-based reporter gene Cell growth, cell viability Cytochrome P450 inhibition Protein binding Protein production by cells ELISA HIGH-THROUGHPUTSCREENING SCREENING HIGH-THROUGHPUT Examples of Assays: Results from Enzyme Inhibition Screen 11,500 compounds screened Normalized results shown Each tile shows 1 plate of data Right: Color Scale for Normalization Distribution of compound % Activity Results from 1 plate in Enzyme Inhibition Screen High Content Screening With the Evotec Opera 96-, 384-, and 1536-well plates Up to ~ 70 images per well in 384-well plate Internalisation Assay Example Image Internalisation Assay Example Image Analysis Internalisation Assay Example Opera External Data output Excel spreadsheet spots / cell cell area nuclei area spots per cell 0.7 0.6 0.5 0.4 spots per cell 0.3 0.2 0.1 0 1 3 5 7 9 11 13 15 17 19 21 23 High Throughput & High Content Screening Services Quality Data Experienced Scientists State-of-the Art Equipment Contacts Sandra Nelson • • • High Throughput & High Content Screening [email protected] 558-2372 Ruben Papoian • • • Drug Discovery [email protected] 558-0516 Charles McOsker • • • Business Development & External Relations [email protected] 558-2569