The High-throughput Screening (HTS) core provides the Perelman School of Medicine community with professional HTS screening services to identify genes or organic small molecule modulators of signaling pathways, cellular phenotypes, and protein function in models of human disease. Core staff will educate and assist scientists with HTS assay development, optimization, miniaturization, and validation; maintain libraries of siRNA, shRNA, cDNA, and FDA approved/FDA-like organic small molecule libraries for HTS; and provide robotics infrastructure and technically trained staff for HTS, including small screens of user defined libraries. The High-throughput Screening core also provides direct assistance with preparation of grant applications by drafting experimental designs approaches and providing Letters of Support, offering HTS resources and analysis expertise for the proposed research.
biosafety level 2 tissue culture
The PerkinElmer EnVision Xcite is a multi-mode plate reader
capable of measuring standard detection labels in 96-, 384-, and 1536-well plates, including absorbance, fluorescence intensity, luminescence, ultra-luminescence, time-resolved fluorescence, fluorescence polarization, and AlphaScreen (Amplified Luminescence Proximity Homogeneous Assay).
• Integrates liquid handling with rapid whole plate imaging of
fluorescence and luminescence
• Can read at sub-second intervals, which enables the kinetics of
the response to be captured
• Integrated pipettor enables successive liquid additions,
providing an opportunity to detect agonists, antagonists, and
allosteric modulators all in one assay
1. Intracellular Ca2+ flux: cell permeable Ca2+ sensitive fluorescent dye (e.g. cardiotoxicity);
2. Fatty acid or neurotransmitter uptake of fluorescence dyes.
3. Membrane potential: lipophilic, anionic, fluorescent dye that partitions across the cytoplasmic membrane of live cells based membrane potential.
Enables automated acquisition of multi-channel
fluorescence images in microtiter plates (5 colors)
Phenotypic assays up to 40X
• Physical properties of cells (size, shape, etc.)
• Puncta formation (autophagy, lipid droplets, etc)
• Activity (migration, invasion, neurite outgrowth)
• Expression and localization of native proteins
• Measurement of fluorescent labels (antibody stains,
EdU/BrdU, Calcein, PI uptake, mitochondria, etc.) • FRET
Matrix Wellmates with integrated stackers and a ThermoFisher Multidrop combi for bulk reagent dispensing. These instruments can be programmed to fill specific plate columns as desired by the user, and is capable of dispensing 0.5-2500 μl per well. The
with 96/384 Modular Dispensing Tool (MDT) and Verispan 8-tip for automated liquid pipetting
"Chembridge (32,000 total): 20,000 from Core set and 12,000 from Express Pick set
ChemDiv (12,000 total): SMART library"
"• Started with ~200,000 compounds
• Used modified Lipinski parameters to filter the set.
• Removed Reactive groups (e.g. Michael acceptors) and compounds with undesirable functionalities.
• Duplicates were removed.
• Lead Finder Clustering and MACCS fingerprints were generated.
• PAINS1, PAINS2 and PAINS3 substructure filters were applied.
• REOS filters were applied.
• Property Based Selection of 50K cpds was performed
• Vendor re-supply validated"
• 1,280 pharmacologically active compounds
• Marketed drugs and pharmaceutically relevant structures annotated with biological activities"
• 18,000 full length, sequenced, mouse and human cDNAs pre-cloned into pCMV-SPORT6
• Insert is fully sequenced and guaranteed to match corresponding BC Accession Number
• Expression-ready vectors eliminate additional cloning steps
• Robust CMV promoters drive cDNA expression
• Gateway sites flanking coding sequence allow for additional flexibility
• Stored as bacterial glycerol stocks, arrayed in 96 well microtiter plates
"The NatProd Collection provides 800 pure natural products and their derivatives, which include simple and complex oxygen heterocycles, alkaloids, sequiterpenes, diterpenes, pentacyclic triterpenes, sterols, many other diverse representatives. The collection is available in 96 well microplates as10mM DMSO solutions or as 5mg solid samples in glass vials."
Screen a library of siRNAs against
• lncRNAs (human, 2220 targets)
• miRNA mimics/antagonists (human V20, 25550 targets each)
Kinase Inhibitors, Epigenetic Inhibitors, Cancer compounds, GPCR/Ion Channel, Metabolism, Microbiology, FDA approved/FDA-like
Human TRC 2.0 and Mouse TRC1.0 Lentivirus shRNA library
• Screening pools: GO categories; user-defined sets
• Order groups/individuals
• human genome-wide
• human GO categories
• user-defined human and mouse
Biochemical, cell and high-content assay development is available.
Assay development involves consultation, technology assessment, assay design, optimization, and miniaturization.
"• Custom siRNA libraries are provided by Ambion/Life
• Minimum order of 20 siRNAs REQUIRED
• 0.1 nmol of Silencer Select siRNA (human)
• 1 nmol of Silencer siRNAs (human and mouse)
• siRNAs are received in microtiter plates (96 or 384)
• All orders are submitted through the HTSC"
• Normalization, annotation
• HCA analysis sequence dev.
• Screen reports
• pharmacologically active compounds
• diversity collections
• focused libraries (e.g. annotated inhibitors)
• siRNA, cDNA, and shRNA
• library plates
with or without help
• letters of support
• experimental design section
"Distribution of Individual clones
• $100.00 per Gene Target (5+ clones)
• Investigator should sequence confirm shRNA sequence
• Investigator responsible for validating knock-down
Preparation of Custom Gene Sets
• Cherry-pick and array clones of interest into new plates ‒ GO categories (e.g. Epigenetic targets) ‒ User-defined sets (e.g. ‘omics validation)
• Glycerol stock plate + pooled plasmid DNA
• 150 gene Targets/625 clones: $2000
• 600 gene Targets/3000 clones: $5000"
• user-defined sets of genes
• user-defined cell-types across small libraries of bioactive compounds/inhibitors
• synergy screening with smaller libraries
• library plates (e.g. kinome)
• Easy analysis pipelines in-house
• Computational analysis of images enables measurement of multiple cellular properties of interest at the object/cell level and/or population level.