NIH Chemical Genomics Center Assay Guidance Manual Table of Contents
From LabAutopedia
| Content transcluded from a collaborative wiki site |
Table of Contents
From The NIH Chemical Genomics Center Assay Guidance Manual
This document is written to provide guidance to investigators that are interested in developing assays useful for the evaluation of compound collections to identify chemical probes that modulate the activity of biological targets. Originally written as a guide for therapeutic projects teams within a major pharmaceutical company, this manual has been adapted to provide guidelines for: Identifying potential assay formats compatible with High Throughput Screen (HTS), and Structure Activity Relationship (SAR) Developing optimal assay reagents Optimizing assay protocol with respect to sensitivity, dynamic range, signal intensity and stability Adopting screening assays to automation and scale up in microtiter plate formats Statistical validation of the assay performance parameters Secondary follow up assays for chemical probe validation and SAR refinement Data standards to be followed in reporting screening and SAR assay results
Authors of Version 6
Assay Validation
- Overview
- Stability and Process Studies
- Plate Uniformity and Signal Variability Assessment
- Replicate-Experiment Study
- How to Deal With High Assay Variability
- Bridging Studies for Assay Upgrades and Minor Changes
- References
Assay Operations for SAR Support
Enzyme Assays
- Basic Enzyme Assays
- Overview of MOA in Drug Discovery
- Glossary of MOA Terms
- Types of Inhibition
- Performing MOA Studies
- Enzyme MOA References
Receptor Binding Assays
- Steps to Assay Development for SPA Format
- Steps to Assay Development for Filter Format
- Introduction
- Reagents
- Scintillation Proximity Assays (SPA)
- SPA Assay Format
- Assay Buffer
- Assay Conditions
- Binding Parameters
- Filtration Assays
- Filter Assay Format
- Assay Buffer
- Assay Conditions
- Binding Parameters
- Special Circumstances
- Practical Use of Fluorescence Polarization in Competitive Receptor Binding Assays
- Abbreviations
- Additional Resources
GTPγS Binding Assays
- Steps to Assay Development
- Introduction
- Materials and Reagents
- Membrane Preperations and Assay Buffers
- Basic Assay Protocol
- Assay Optimization
- Data Analysis
- Filtration Assays
- Non-Radiometric GTPγS Assays
- Literature References
Tissue Culture Assays
- Cell Culture Glossary
- Introduction to Cell Based Assays
- Requirements for Assay Proof of Concept for Cell Based Assays
- Tissue Culture Assay Development, Optimization and Validation Flow Chart
- Types of Tissue Culture Assays
- Batch Co-Transfection and Cell Banking Protocols
- Cryopreservation of Cells
- Cell Counting Protocols
- Factors for Cell Based Assay Development
- Typical Cell Culture Protocol
- Standard HTS Cell Culture Practices to Reduce Contamination Risk
- Example, Transient Transfection of Cells
- Transient Transfection Methods
- Transfection Cationic Matrix Reagents
- General Cell Biology Information
- References
Cell-Based Elisa (C-Elisa) and Westerns Blots for Quantitative Antigen Detection
- Introduction
- Cell Culture and Treatments
- Fixing and Staining
- Reagents
- Assay Validation
- Appendix 1. Immunoblot vs. Cytoblot Assays for Enzyme Inhibitor Potencies
- Appendix 2. Linearity and Sensitivity of Cell-Based Western Assays
- References
Additional Functional Assays for GPCRs and Ion Channels
- cAMP Detection
- β-Arrestin Recruitment
- IP-3/IP-1 Assays
- Impedance-Based Technologies
- Phospho-ERK Assays
Flipr™ Assays to Measure GPCR and Ion Channel Targets
- Overview: Flipr™ Assay Development
- Introduction
- Types of Flipr™ formats
- Reagents and Buffers for Method Development
- Method Development and Optimization
- Flipr™ Instrument Set-Up
- Potential Artifacts
- References
Immunoassay Methods
- Immunoassay Development, Optimization and Validation Flow Chart
- Introduction
- Immunoassay Parameters
- Reagents
- Instrumentation
- Immunoassay Formats
- Important Parameters for Development of an Immunoassay
- Initial Concept and Method Development of a Sandwich Immunoassay
- Results
- Second Experiment-Matrix Compatibility
- Initial Concept and Method Development of a Competitive Assay
- Development of a Competitive Immunoassay
- Method Validation (Pre-Study)
- Method Validation (In-Study)
- Pre-Study & In-Study Acceptance Criteria
- References
Guidelines for Cell Proliferation Assays
- Cell Proliferation Assays
- Methods for Evaluating Cell Proliferation
- Setting Up a Cell Proliferation Assay
Cell-Based RNAi Assays
- Introduction
- RNAi Loss-of-Function Screens
- RNAi Synthetic Lethality Screens
- Appendix
- Acknowledgements
- References
- Abbreviations
Data Standardization for Results Management
- Introduction
- Data Types and Associated Rules for Radioligand Binding Assays: Inhibition Mode
- Data Types and Associated Rules for Enzymatic Assays: Inhibition Mode
- Data Types and Associated Rules for In Vitro Functional Assays
- Glossary
Calculations and Instrumentation Used for Radioligand Binding Assays
- Introduction
- Radioactive Calculations
- Instrumentation
- Uniformity Plate
- Color Quench Correction
- Abbreviations
Assay Equipment and Instrumentation
- Introduction
- Common Equipment in HTS Labs
- Spectrophotometry
- pH meters
- Electronic Balances
- Microscopes
- References
In Vivo Assay Guidelines
Glossary of Quantitative Biology Terms
Sites Collaborating with NCGC
----------------------------------------------------------------------------------------------------
Trial Sections- To be reviewed and approved by Editorial Board
Section 19 - Stem Cells, Regenerative Medicine & Chemical Biology -G. Sitta Sittampalam
| Click [+] for other articles on | The Market Place for Lab Automation & Screening | Drug Discovery HTS Products |
