Life Sciences WNT PATHWAY. Integrated solutions for screening Wnt regulators. Image: TR Arnett, UCL

Transcription

1 Life Sciences WNT PATHWAY Integrated solutions for screening regulators Image: TR Arnett, UCL

2 3Find Your Total Solution

3 SOLUTIONS FOR WNT PATHWAY SCREENING High-throughput Assays for Canonical Pathway Analysis Since the discovery of the gene over 30 years ago as the Int1 proto-oncogene involved in the pathology of Mouse Mammary Tumor Virus (MMTV), expression and signal transduction have been identified as critical components in regulating embryogenesis, cell proliferation, and bone, glucose, and lipid metabolism. Perturbations in expression, surface receptors, signaling partners (including β-catenin and GSK-3β), and the battery of -regulated genes (collectively the canonical pathway) are known to manifest in a variety of clinically relevant disease states, including cancer, osteoporosis, cardiovascular disease, diabetes, and neurodegeneration. WNT MODULATOR SCREENING WIF-1 PATHWAY ANALYSIS Screen-Well Pathway Library SOST DKK Kremen sfrp Proteasomal degradation Axin Frat-1 Dsh ELISAs Dkk-1, GSK-3β, [pser9] GSK-3β, β-catenin Leading Light TM Reporter Assay APC Axin P Nuclear Localization p300/cbp Regulated Gene Expression TCF/LEF Antibodies GSK-3β, GSK-3 α/β, Dishevelled-1, Dishevelled-2, β-catenin Proteins APC, Dkk-1, GSK-3β, -3a, Casein Kinase II Inhibition Receptor Activation β-catenin phosphorylation by β β-catenin degredation Activation Receptor Activation β-catenin phosphorylation by β β-catenin nuclear localization Cellular Analysis We have extensive experience in building next generation cell based assays for use in drug development, secondary screening, and pathway analysis. Our Leading Light and CELLestial assay systems are optimized to meet the needs of demanding highthroughput applications to deliver better, high-quality hits. Our portfolio of over 300 ELISA kits includes sensitive, specific assays for key signaling proteins in the canonical pathway in a variety of species. In addition, our catalog of over 3000 antibodies and recombinant proteins includes thoroughly validated reagents for dissection of pathway signaling. Small Molecule Chemistry Our long-standing, flagship Screen-Well Compound Library portfolio offers an easy, ready-to-use method to streamline compound screening. Included within our thousands of stand-alone small molecules is a unique collection of Pathway targeting compounds as well as focused libraries for kinase inhibitors, natural products, protease inhibitors, nuclear receptors, autophagy regulators, and more. Through our supplier network built over a 25 year history, we have the ability to rapidly and inexpensively source traditional, custom, and bulk compounds in the gram to kilogram scale. We complete our sourcing using stringent quality control standards with state-of-the-art methods.

4 SCREEN WNT PATHWAY ACTIVATORS & INHIBITORS Development of pathway modulating drugs requires the ability to rapidly screen the effect of novel or repurposed compounds and biological therapeutics on the expression of -regulated genes, and confirmation of such effects by related changes in key signaling components. Together, the Leading Light Reporter Assay and Screen-Well Pathway Library facilitate high-throughput analysis of -related signaling. Leading Light Reporter Assay (ENZ-61001) Reliable, All-inclusive Screening of Pathway Modulators The Leading Light Reporter Assay Starter Kit is a high-sensitivity, luciferase-based assay for the analysis of canonical signaling. The assay uses a 3T3 mouse fibroblast cell line modified with a luciferase reporter gene under the control of -responsive promoters (TCF/LEF). The chemiluminescent readout is amenable to both 96-well and 384-well plate formats, facilitating high-throughput analysis of pathway activators and inhibitors. Screen your Pathway in only 4 steps! All kit components included! BENEFITS A true end-point detection system for screening compounds Greater sensitivity than other whole cell assay kits on the market Reproducibility that comes from using a validated system Time-saving, no-transfection kit includes cells, media, detection reagents and controls 1. Plate 2. Treat 3. Develop 4. Detect Leading Light TM Cells Growth Medium Assay Medium Freezing Medium -3a Protein Dkk-1 Protein Lithium Chloride (Optional) Luciferase Substrate Luciferase Assay Buffer Activation by -3a Protein (Activator) (Activator) Dsh Frat-1Nuclear Localization In-tact Pathway For Screening Flexibility - Small Molecules - Biotherapeutics - Genetically Modified Cells (e.g. transfect, sirna, etc.) Axin Luciferase Luciferase Luciferase p300/cbp Luciferase Reporter TCF/LEF Leading Light TM Inhibition by Dkk-1 TCF TCF TCF TCF TCF TCF TATA FIREFLY LUCIFERASE Reporter Cell Line (Activator) + Dkk-1 (Inhibitor) WIF-1 sfrp SOST Proteasomal degradation Dkk Kremen APC P Axin Pathway Compound Library (BML-2838) The Screen-Well Pathway Library offers a convenient solution for benchmarking compounds against a collection of 75 known modulators. The library is available in 100uL and 500uL formats (dissolved in DMSO) for use as a companion to the Leading Light Reporter Assay, or individually as a general screening tool for chemical genomics or assay. ScreenWell TM Pathway Library Targets Include 5a Dkk Axin Porcupine β-catenin TCF Dishevelled GSK-3B Adenylate Cyclase COX-II sfrp-1

5 QUANTIFY CRITICAL WNT PATHWAY PROTEINS Sensitive Pathway ELISA Kits Obtain fully quantitative data with immunometric detection of, total and phosphorylated GSK-3β, and DKK-1 in human, mouse and rat samples. Sensitive assays measure picogram levels compared to microgram levels in Western blot Suitable for cell lysates, culture supernates, plasma, or serum sample types (see table) High throughput format allows analysis of 40 samples in duplicate in less than 3 hours Simple protocols and liquid color-coded reagents save time and reduce error PRODUCT SENSITIVITY SPECIFICITY SAMPLE TYPES Dkk-1 Human Mouse Rat Cell Lysates Culture Supernates Serum Plasma ADI pg/ml ADI pg/ml ADI pg/ml GSK-3β Human Mouse Rat Cell Lysates Culture Supernates Serum Plasma ADI pg/ml [pser9] GSK-3β Human Mouse Rat Cell Lysates Culture Supernates Serum Plasma ADI A 9.0 pg/ml β-catenin Human Mouse Rat Cell Lysates Culture Supernates Serum Plasma ADI pg/ml Highly Characterized Related Proteins & Antibodies Enzo offers a wide selection of high-quality reagents for analysis of related signal transduction, including purified proteins and thoroughly validated monoclonal and polyclonal antibodies. Select Pathway Proteins & Antibodies Product Name Product # Product Name Product # Casein Kinase II (human), (recombinant) BML-SE124 Adenomatous polyposis coli (APC), pab ADI Dkk-1 (mouse), (recombinant) (His-tag) ADI-KPR-CP100, mab (12F7) ADI-KAM-ST001 GSK-3β (human), (recombinant) (GST-tag) BML-SE355 Dishevelled-1 pab BML-DA4170-3a (mouse), (recombinant) ENZ Dishevelled-2 pab BML-DA4270 GSK-3α/β, mab (1H8) ADI-KAM-ST002 GSK-3β, pab ADI-KAP-ST002 Monitor signal transduction from cell surface to nucleus.

6 Life Sciences scientists enabling scientists. GLOBAL HEADQUARTERS Enzo Life Sciences Inc. 10 Executive Boulevard Farmingdale, NY Phone: Fax: EUROPE/ASIA Enzo Life Sciences (ELS) AG Industriestrasse 17 CH-4415 Lausen, Switzerland Phone: Fax: For local distributors and detailed product information visit us online: MB Put our experience to work for you! Contact Enzo to learn more about how our scientists are working to provide innovative tools for your research.