The Small Molecule Screening Facility (SMSF) has specialized equipment dedicated to drug discovery. Scientists from UW-Madison, other universities or private companies may receive training on some pieces of the shared equipment and then use the online appointment system to schedule “walk-up” use.
If you have questions or problems with this system, please contact Tom Polaske. Appointments for equipment training can be made by contacting Tom Polaske (polaske@wisc.edu) or Song Guo (sguo6@wisc.edu)
Notice: The SMSF is undergoing rates evaluation after a 5+ year price freeze. We will be implementing a comprehensive 5% equipment and personnel rate increase on February 1, 2024.
High-throughput screening (HTS) combines robotic liquid handling and assay miniaturization (HTS is typically performed in 96-well, 384-well, and 1536-well microtiter plates) with data analysis software and computational methods to identify hit compounds and their families. Users can leverage our HTS instrumentation and expertise to screen libraries of over 500,000 “drug-like” small molecules, 300,000 natural product fractions, or 18,000 human siRNAs. The experienced SMSF staff are available to assist in assay development, assay validation, assay implementation, data analysis and chemical informatics.
The first step in initiating a high-throughput screen at the SMSF is to contact facility manager Dr. Tom Polaske to make an appointment to discuss your project and goals. There is no charge for these initial discussions. In addition to primary HTS assays, it is typically prudent to identify secondary assays to confirm hits and counterscreens to eliminate false positives. We strongly encourage, but do not require, a student, postdoc or technician from the client lab to assist “hands-on” with the HTS at SMSF.
A frequently asked question is “How much will it cost?” This depends on the specifics of the assay, availability of reagents from the client lab, and the number of compounds or RNAi pools screened. Generally HTS assays cost from $0.10 to $1.00 per well, including reagents and the time of the SMSF scientists. Every effort will be made to provide the client with a realistic estimate of costs prior to beginning a project. This is an estimate and not a contract price.
The greatest uncertainty in predicting the cost of a project is the staff time required during the initial phases of development (optimization) of a client assay. That is, it is cost efficient (but not necessary) for clients to already have an optimized assay (or at least have progress made to this end) before starting a HTS campaign. A properly-optimized assay will have considered plate format, readout(s), controls, environmental conditions, experimental compound concentration(s), and statistical performance. Details, concerns, and questions regarding assay optimization can and will be discussed during the initial free consultation. Charges for time and supplies will be billed on a frequent, usually monthly, basis thereby allowing the client to decide whether a project should be continued. It is our goal to deliver the best results possible while eliminating extraneous expenses for clients.
The in-house compound collection includes over 500,000 small molecules and 300,000 natural product fractions, but most clients will only screen a subset of the compound libraries. The number of compounds screened is a key variable in determining the overall costs of a project and is often dependent on the goal of the project. Radioactive assays are currently not supported by SMSF and substances requiring a higher containment level than BSL2 are not permitted.
A terrific resource on HTS assays is the NIH-supported Assay Guidance Manual. Signal/background ratio (S/B) and well-to-well variability (CV) are key variables in HTS. See the HTS Assay Validation Chapter in the Assay Guidance Manual for a discussion of how the statistical robustness of an HTS assay will be determined by SMSF scientists.
Once an assay is demonstrated to be statistically robust, a small pilot screen of 100 – 1,000 compounds may be run in duplicate plates to confirm assay reproducibility and assess the hit rate. The client may then make a decision as to how many compounds to screen from the in-house collection.
SMSF scientists are available to assist in analysis of the screening data. After completing a primary HTS, clients typically will have several screening positives or “hits” to follow-up in secondary screens. Subject to compound availability, SMSF will provide clients with 1 µl of each selected screening hit (~5 µg). These are called “cherry picks.” If you require more than 1 µl of compound, SMSF stay may assist you in ordering it directly from a commercial source. Generally, 1-5 mg of a compound can be purchased around $25 to $100 from the chemical manufacturers.
In addition to secondary assays, we recommend SMSF services for hit identification and triage, to evaluate the compounds, and assist in planning the follow-up procedures.
GlaxoSmithKline (GSK) Protein Kinase Inhibitor Set
Received July 2014, the Protein Kinase Inhibitor Set (PKIS1 and PKIS2) consists of 868 small molecule kinase inhibitors that have been published by GSK
Representing a range of chemotypes as well as multiple exemplars within a given chemotype
Broad coverage of kinase targets, characterized across a panel of 220 kinase assays.
NCI
Received October 2012
The NCI04 “Diversity Set III” library consists of 1597 compounds.
The NCI05 “Mechanistic Diversity Set” library consists of 879 compounds which have represent a broad range of structural diversity and patterns of growth inhibition in the NCI 60 cell line screen.
The NCI06 “Natural Product Set II” library consists of 117 natural products that were selected from the NCI open repository.
The NCI07 “Approved Oncology Set IV” library includes 101 compounds.
Received July 2014, the Anti-Infectives Library includes 960 molecules and contains low molecular weight, drug-like molecules with scaffolds found in antiseptic agents with anti-bacterial (Gram+ve and Gram-ve), anti-fungoid, anti-microbial activities.
Life Chemicals
(received #1- January 2009, #2- October 2009, #3- November 2015, #4 – October 2016)
The Life Chemicals diversity libraries (LC-1, LC-2, LC-3, LC-4) total 100,000 diverse compounds, arrayed in subsets of 25,000. Each set was selected for diverse coverage of chemical space and general drug-likeness. Available for screening as each subset or the full library.
Maybridge
(received October 2008)
The Maybridge HitFinder collection consists of 14,400 compounds representing the drug-like diversity of the Maybridge compound catalog.
MLPCN: NIH Molecular Libraries Probe Production Centers Network Diversity Library
(received May 2014)
The MLPCN library consists of 340,364 small molecule probes, covering a wide range of chemical space.
NCI NExT Diversity Library
(requested as needed from NCI for oncology-related HTS campaigns that received funding through a peer-viewed process) The set contains a variety of chemical scaffolds across ten commercial suppliers, while keeping within the boundaries of drug-like chemical space (e.g. Lipinski’s Rule of 5, high QED scores). The collection includes a total of 83,536 compounds.
The NCI Program for Natural Product Discovery (NPNPD) Prefractionated Library
The NPNPD Prefractionated Library is a collection of >295,000 partially-purified natural product fractions available for use in high-throughput screening.
Important source of new drugs and drug leads
Comprised of plant, marine invertebrate, and microbial samples
Hit fractions can be selected for further purification by the NCI at no cost
Tip-less acoustic liquid handling using the Labcyte ECHO can transfer as little as 2.5nL of DMSO compound solution between plates. This capability allows us to offer assay ready plates for screening at low cost. The resulting plates are ready to accept reagents for screening in your lab or ours. Control wells can be formatted as desired. The SMSF Diversity and Specialized libraries are available in 96-well, 384-well and 1536-well formats in a wide range of plate styles. For additional details or prices, please contact Dr. Tom Polaske (polaske@wisc.edu).
The Small Molecule Screening Facility (SMSF) offers a broad range of biological profiling assays to help identify a chemical compound’s biological activity and potency including:
AlphaLisa and AlphaScreen Assays for Biomarkers, Cytokines, Phospho-proteins and more
AKT Phosphorylation
Anti-microbial Assays
Apoptosis Assays
Cell Migration
Cytotoxicity and Cell Proliferation Assays
ERK1/2 Phosphorylation
HDAC
NFKb
Phosphatase Assays (PP1A and PP2A)
Phosphorylation
Reporter Gene Expression
Signal Transduction
TGF-beta signaling
Viral Entry
WNT -Beta Catenin (Super Topflash Assay)
Wound Healing and Chamber Migration Assays
Custom assay development services are also available.
SMSF is the home of a new Seahorse Bioscience XFe96 Extracellular Flux Analyzer which simultaneously interrogates mitochondrial respiration and glycolysis – in a microplate, in real-time. The XFe96 Analyzer determines in vitro oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) in order to assess cellular functions such as oxidative phosphorylation, glycolysis and fatty acid oxidation.
Real-time Results- this integrated system reports metabolic rates in just minutes, without sample extraction or labeling. Wave software controls the instrument and performs rate measurements on the fly for same day results.
Live-Cell Responses – detect responses to substrates, inhibitors, and other compounds in real time via the 4-port injection system and automated mixing while maintaining physiologic temperature (37°C).
Flexible Assay Design- the 96-well plate format accommodates many conditions in a single run and is best for dose-response studies and compound screening.
High Sensitivity- analyze as few as 5000 cells per well in the custom 96-well plate. Cell number requirements vary by cell type; consult the Cell Reference Database for details.
Compatible with a variety of samples due to the precision-controlled heating tray, which can maintain 16-42oC (12oC-20oC above ambient conditions).
Measure mitochondrial function with the Seahorse XF Cell Mito Stress Test
Generate a metabolic phenotype within one hour with the Seahorse XF Cell Energy Phenotype Test
Assess glycolytic rates in live cells with the Seahorse XF Glycolytic Rate Assay
Quickly determine dependency of cellular energy production on mitochondrial substrates with the Seahorse XF Mito Fuel Flex Test
Analyze 3D samples such as spheroids and islets with the Seahorse XFe96 Spheroid Plate
Easily create assay protocols and analyze data with Seahorse Wave software
For Research Use Only. Not for use in diagnostic procedures.
Mycoplasma contamination is one of the most common and serious problems in culturing eukaryotic cells because it alters the phenotypic characteristics of the cells and can negatively impact results. Since mycoplasma is typically not visible and does not respond to antibiotics, an alternative sensitive and reliable detection method is required. Small Molecule Screening Facility (SMSF) uses the R&D Systems Mycoprobe Mycoplasma Detection Kit. This assay is capable of detecting the eight mycoplasma species known to cause 95% of eukaryotic cell culture contamination: M. hyorhinis, M. arginini, M. fermentans, M. orale, M. pirum, M. hominis, M. salivarium and Acholeplasma laidlawii.
The SMSF staff scientists test each client-provided sample in triplicate along side positive and negative controls.
Assays are usually run the week that samples are submitted and data are available within 48 hours of assay completion.
The SMSF can provide a variety of molecular modeling services to guide the identification of small molecules that bind to protein targets of interest.
Pymol image of the tgfbeta Smad3/Smad4 complex with mutation sites in spheres and docking sites as translucent surfaces. Smad3 molecules are green and magenta, and smad4 is grey. Parts of the binding proteins are ski (salmon) and sara (blue).
Virtual Screening
Homology Modeling
Docking and Scoring
QSAR Modeling
HTS Data Analysis and Compound Selection
Structure-based or Ligand-based molecular design
Protein Molecular Dynamics
We will work with researchers to determine the methods that are most appropriate for each particular project.
Our virtual screening techniques can be applied to most projects where either a protein target or potent ligand is known. Docking and scoring is most commonly applied in conjunction with synthetic chemistry effort.
We are actively involved in optimization of methods for virtual screening and machine learning approaches for drug discovery. We collaborate with investigators from Biostatistics, Computer Science, Engineering, Biochemistry, and the Center for Predictive Computational Phenotyping on these efforts, and use these novel techniques on our discovery projects.
Our computation resources include 5 Dell servers and workstations with a total of 48 CPUs, including NVIDIA 3D graphics capabilities. We make extensive use of computational resources at the UW Center for High Throughput Computing (CHTC) through HTCondor. Common software includes Autodock (Scripps), Vina (Scripps), ROCS and FRED (OpenEye), Sybyl (Tripos), Surflex (UCSF), MOE (CCG). An additional workstation is available for researchers who wish to actively participate under the supervision of our team.
Hands-on Course in Molecular Modeling and Drug Docking
The SMSF also sponsors a “hands on” 8-week course on molecular modeling with an emphasis on drug design called “Molecular Modeling and Drug Docking” for the UW community to learn how to model the drugs in their targets. The course covers the basics of protein and small molecule modeling using the commercial software Sybyl from Tripos. Several docking programs, such as SurFlex, DOCK and Autodock4, are examined with real examples from the literature.
The course is held in the Digital Media Center teaching lab in the Biochemical Sciences Building, 420 Henry Mall. Each student runs Sybyl on a Linux server thru a Macintosh interface. There is no UW credit for this course. The course has been taught the last five years by Dr. Ken Satyshur from the SMSF during the Spring semester.
Preclinical in vitro Pharmacology Assays
DDC staff can assist users with characterizing the pharmacological properties of compounds through contracts maintained with external CROs.
Five test compounds per mouse, 1dose, 1 route, 0.5hr and 2hr
Five test compounds per mouse, 1dose, 1 route, 0.5 h, 2h, 4h, and 7h
Single Dose Tolerability (1 day), Five dose levels, 1 route, 7 Blood sampling times up to 24h post dosing per animal
Multi- Dose Tolerability (5 days), Five dose levels, 1 route, Animals will receive repeat dose once daily for five days, 7 Blood sampling times up to 24h post dosing per animal
Equipment
HTS and Profiling Assay Equipment
Training and Scheduled Walk-up Use Available
$84/hour base charge; minimum time 15 minutes
Liquid Handling (available in Biosafety Cabinet if sterile transfers needed)
Formulatrix Tempest contact-free dispenser
BlueCatBio Blue Washer contact-free plate washer for 96, 384 and 1536 well plates
Sorenson Benchtop Pipetter 96 and Benchtop Pipetter 96-low volume
Biotek Microflo peristaltic reagent dispensers
ThermoFisher Multidrop Combi
Plate Readers and Scanners
BMG Pherastar Multimode plate reader for absorbance, luminescence, AlphaELISA, and filter-based fluorescence and fluorescence polarization
BMG Clariostar plate reader for absorbance, luminescence, and fluorescence with monochromator
Walk-up Use Not Available
Beckman fX Liquid Handling (96 and 384)
Labcyte Echo 650
Seahorse Metabolism Analysis
Seahorse XFe96
Pricing
The Small Molecule Screening Facility (SMSF) operates on a fee-for-service model. User fees pay for staff scientist salaries, consumables and reagents. UWCCC Members receive subsidized rates.
The broad range of services and the individually tailored projects preclude a simple fee schedule. To obtain an estimate for your specific project, please contact us to discuss your needs. There is no charge for meetings or conference calls to discuss project details. Staff scientists will provide an estimate for the project; this is not a contract price, but a conservative estimate based on experience. Projects may include intermediate evaluations of progress and costs to permit clients to make “go/no go” decisions about continuing a project, especially in the event of unexpected complications that increase the cost of the project.
Before SMSF initiates a project, the client must provide a UW fund number, or a purchase order number for off-campus clients. Costs will be billed monthly. Detailed invoices showing time logged by staff scientists on the project and all supply costs are prepared monthly and available to the client or their departmental administrator.
