A Guide to Assay Development

A Guide to Assay Development

Click to purchase: A Guide to Assay Development

Description: D&MD's Guide to Assay Development provides critical information to speed and simplify the development and optimization of most technologies. This Guide eliminates the tendency to "reinvent the wheel" by providing valuable tips that incorporate the author's many years of hands-on experience in the field.

300+ Pages, 45+ Exhibits, Including Assay Development Checklists & 2 Examples of Assay Troubleshooting

This Guide thoroughly evaluates the capabilities, strengths, weaknesses, and expectations of leading assays and assay technologies.

Assay development is often time consuming and frustrating. But reliable, robust assays are key to every phase of pharmaceutical development. D&MD's Guide to Assay Development provides critical information to speed and simplify the development and optimization of most technologies. This Guide eliminates the tendency to "reinvent the wheel" by providing valuable tips that incorporate the author's many years of hands-on experience in the field.

This Guide provides the information necessary to select and develop the appropriate assay for the task at hand, including:

The most common and rapidly-growing assays in the drug discovery and drug development industries.
Charts and tables cut through the sales hype and facilitate comparisons of different assay types.
Tips and tricks learned over 15 years of professional experience and collaborative work in laboratory assays.
The often neglected topics of biological sample matrix and high-volume data analysis.
Step-by-step instructions and checklists for assay optimization, validation, automation, scale-up, and miniaturization.
Emerging platforms, formats, and technologies for high-throughput screening (HTS) and ultra-high-throughout screening (UHTS).
Detailed comparisons of technologies designed to help scientists determine the best format to meet their assay needs.
Lists of instrument manufacturers and supply vendors-including original references and historical developments, with resources for finding detailed modern laboratory recipes.

The following professionals involved in drug discovery, screening, lead generation, and assay technology will greatly benefit from this Guide:

Directors, Lab Managers, Group Leaders, Senior Scientists, Principal Scientists, Project Managers, Heads of Research & Development, Quality Control Managers, and more

This Guide Will Answer the Following Questions:

How does a scientist determine the best assay for a particular application?
What questions must be asked and answered during assay development?
What steps are involved in optimization and validation of an assay?
How can a scientist determine that an assay is fully optimized, sufficiently validated, and ready for full-scale operation?
How can controls be used to provide the earliest warning of assay problems?
How does sample throughput and assay miniaturization really affect data quality and reagent consumption?
Will migrating to a higher-density microplate format improve throughput?
How do state-of-the-art assay instruments fail during normal operation, and what can be done to minimize failure and the impact of failure on operations?
What universal considerations are critically important for every assay?

Table of Contents: Executive Summary, Introduction, Common Assays, General Assay Design, Format, Readout (Reporting Format), Validation, In Vitro Assays, Cell-Based Assays, Automation Platforms, Emerging Technologies, Information Management, Appendices, Bibliography

Table of Contents
Section	Summary
1	Executive Summary
2	Introduction
2.1	Definitions
2.1.1	Assays and Analytes
2.1.2	Qualitative and Quantitative
2.1.3	Targets
2.1.4	Stains, Dyes, Tags, Labels, and Reporters
2.1.5	Potentially Confusing Terminology
2.2	Brief Historical Perspective
2.2.1	Diagnostics
2.2.2	Drug Discovery
2.3	Basic Issues
2.3.1	Sampling
2.3.	2 Primary and Secondary Assays
2.3.3	Confounding Analytes
2.4	Drug Development and Manufacturing
2.4.1	Impurities
2.4.2	Therapeutics
2.5	Miniaturization
2.5.1	Microplates
2.5.2	Arrays and Microarrays
3	Common Assays
3.1	Quantitation of Therapeutics and Other Compounds
3.1.1	Weighing
3.1.2	Extinction Coefficient
3.1.3	Evaporative Light Scattering Detection
3.1.4	Nuclear Magnetic Resonance
3.1.5	Chemiluminescent Nitrogen Detection
3.2	Microbial Contamination
3.2.1	Culture Tests
3.2.2	PCR
3.2.3	ELISA
3.3	Binding
3.3.1	Molecular Size
3.3.2	Biochemical Function
3.3.3	Labeling
3.4	Enzyme Assays
3.4.1	Proteases
3.4.2	Kinases and Phosphorylases
3.5	G-Protein Coupled Receptors
3.5.1	Cell Culture Expression
3.5.2	Promiscuous GPCR
3.5.3	cAMP
3.5.4	Calcium and IP3
3.6	Ion Channels
3.6.1	Ion Flux
3.6.2	Patch-Clamp
3.6.3	Reporter Dyes
3.6.4	Voltage Sensitive Dye Systems
3.6.5	Membrane Binding Assays
3.7	Toxicology and Pharmacology
3.7.1	Ion Channel Assays in Cardiac Toxicity
3.7.2	Gene Expression in ADME
3.7.3	Hepatotoxicity
3.8	Genetic Polymorphism
3.8.1	Restriction Fragment Length Polymorphisms
3.8.2	Hybridization Assays
3.8.3	PCR for SNP
3.8.4	Single Base Extension
4	General Assay Design
4.1	Universal Considerations
4.1.1	Precision 1
4.1.2	Optimal Reagent Amounts
4.1.3	Standard Curves
4.1.4	Multiplicity and Statistics
4.1.5	Interpolation
4.2	Nature of Analyte and Matrix
4.2.1	Analyte Stability
4.2.2	Separation and Enrichment
4.2.3	Internal Standard "Spike"
4.3	Assay Objectives
4.3.1	Research, Development, or Process
4.3.2	Budget
4.3.3	Throughput
4.3.4	Scalability
4.3.5	Sensitivity
4.3.6	Error Tolerance
5	Format
5.1	Homogenous and Heterogeneous
5.1.1	Pros and Cons
5.1.2	Heterogeneous Immobilization
5.2	Direct and Indirect
5.2.1	Second Mediators
5.2.2	Second Antibodies
5.2.3	Biotin-Avidin
5.2.4	Enzyme Reporter Systems
5.3	Agonists and Antagonists
5.3.1	Competition
5.4	In Vitro and In Vivo
5.4.1	In Vitro Assays
5.4.2	In Vivo Assays
5.4.3	Biological Material
6	Readout (Reporting Format)
6.1	Colorimetric and Fluorometric
6.1.1	Colorimetric Assays
6.1.2	Fluorometric Assays
6.2	Radiometric
6.2.1	Radiolabeling
6.2.2	Radioimmunoassay
6.2.3	Scintillation Proximity Assay
6.3	Biological Growth
6.3.1	Special Growth Media
6.4	Other Readouts
7	Validation
7.1	Installation and Operation Qualification
7.1.1	Instruments and Equipment
7.1.2	Assay Optimization
7.1.3	Other Variables
7.2	Performance Qualification
7.2.1	Instruments and Consumables
7.2.2	Operation Re-Certification
7.2.3	Assay Parameters
7.2.4	Data Certification
7.3	Scale-Up
7.3.1	General
7.3.2	Step-by-Step Process
8	In Vitro Assays
8.1	General
8.1.1	Spectroscopy
8.1.2	Protein Assays
8.2	Enzyme Assays
8.2.1	Enzyme Stability
8.2.2	Proteases
8.2.3	Kinases and Phosphorylases
8.2.4	Enzyme SPA
8.3	Binding
8.3.1	Specificity
8.3.2	Valence and Avidity
8.3.3	Blocking and Washing
8.4	Antibodies and Immunoassays
8.4.1	Antibodies
8.4.2	Immunoassays
8.4.3	ELISAs
8.4.4	"Classical" Immunoassays
8.4.5	Fluorescence Quenching
8.4.6	Fluorescence Polarization
8.4.7	Scintillation Proximity Assay
8.4.8	Other Binding Assays
8.5	Nucleic Acids
8.5.1	General
8.5.2	Polymerase Chain Reaction
8.5.3	Hybridization Blots
9	Cell-Based Assays
9.1	General
9.1.1	Pitfalls
9.1.2	Controls
9.2	Fixed Cells
9.2.1	Fixing
9.2.2	Fluorescence in Situ Hybridization
9.2.3	Immunofluorescence Assay
9.2.4	Staining and Counter-Staining
9.3	Whole Cells
9.3.1	Cell ELISAs and ELISpots
9.3.2	Cytotoxicity
9.3.3	Surface Binding and Membrane Transport
9.3.4	Trafficking and Translocation
9.3.5	Nuclear Receptors
9.3.6	Cell Quantitation
9.3.7	Vitality
9.3.8	Motility
9.3.9	Apoptosis
9.3.10	SPA in Microplates
9.3.11	Fluorescence
9.4	Flow Cytometry
9.4.1	General
9.4.2	Advantages and Limitations
9.4.3	Fluorophore Selection
9.4.4	Controls
9.4.5	Gating
9.4.6	Applications
9.5	Lysates
9.5.1	Gene Expression
9.5.2	Protein Expression (Gene Induction)
9.6	Microscopy
9.6.1	Stains
9.6.2	Labels
9.6.3	FISH and IFA
10	Automation Platforms
10.1	Miniaturization
10.1.1	Pros and Cons
10.1.2	Scaling
10.2	Robotics
10.2.1	Conveyors and Workstations
10.2.2	Integrated and Modular
10.2.3	Examples
10.3	Liquid Handlers
10.3.1	General Applications
10.3.2	Delivery Size
10.3.3	Calibration
10.3.4	Cross-Contamination
10.3.5	Pipet Tips
10.4	Cell Handling
10.4.1	Culture
10.4.2	Sample Prep
10.5	Microplate Equipment
10.5.1	Microplates
10.5.2	Plate Handlers
10.5.3	Washers
10.5.4	Bar Codes
10.5.5	Cleaning and Maintenance
11	Emerging Technologies
11.1	Automated Image Analysis
11.1.1	General Issues
11.1.2	Practical Considerations
11.1.3	An Example
11.2	Higher Density Formats
11.2.1	384-Well Plates
11.2.2	High Density Microplates
11.2.3	Microarrays, Microfluidics, and Chips
11.3	New Technologies
11.3.1	Surface Plasmon Resonance
11.3.2	Flow Cytometry with Labeled Beads and Libraries
11.3.3	Branched DNA Binding Assay
11.3.4	Single Molecule Detection
11.3.5	Virtual Screening
12	Information Management
12.1	Data Analysis
12.1.1	Acquisition
12.1.2	Signal-To-Background and Signal-To-Noise Ratios
12.1.3	Precision and Accuracy
12.1.4	Type 1 and Type 2 Error
12.1.5	Random and Systematic Error
12.1.6	Binning and Pooling
12.2	Statistics
12.2.1	Binding Constants
12.2.2	Michaelis-Menten Equations
12.2.3	Error and Standard Error
12.2.4	Correlation Coefficient
12.2.5	Standard Deviation
12.2.6	Coefficient of Variance
12.2.7	Z' Factor
12.2.8	More Complicated Statistics
13	Appendices
13.1	Resources for Detailed Protocols
13.1.1	Orgs
13.1.2	Journals and Other Commercial Publications
13.2	Checklist for General Assay Development.
13.3	Microarray Assay Checklist
13.4	Vendors
13.5	Examples of Troubleshooting
13.5.1	In Vitro
13.5.2	In Vivo
14	Bibliography

Date published: 3/1/2003
Number of pages: 300+
Category: Drug Discovery
Industry: Pharmaceutical & Biotech

Price Hard Copy (US$): $395.00

Interested in this report: peter.barfoot@bioportfolio.com

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