Digital PCR Technology Report

  • June 2013
  • -
  • CHI Insight Pharma Reports

The newly released Insight Pharma Report is about the digital PCR technologies that have surfaced in the market. Digital PCR is a new application in research and development allowing for higher sensitivity and specificity than real-time PCR. This allows researchers to closely examine rare genetic mutations (including single nucleotide polymorphisms), copy number variations, viruses, prenatal defects, and much more. National measurement institutions have also been looking into digital PCR as a reference standard as well as using its precision and accuracy to create reference standards for diagnostic labs.

This report focuses on the main vendors supplying digital PCR systems, validation of the instruments, and digital PCR's impact in research. By detecting rare sequences that were previously undetected with real-time PCR, digital PCR has enabled scientists to accelerate their research. However, there are many areas of the new technology that have been overlooked and neglected, and this report covers those aspects as well.

Also included in this report are:

• Interviews with the main vendors in the market space including
- RainDance Technologies
- Bio-Rad Laboratories
- Life Technologies
- Fluidigm Corporation

• Interviews with national measurement institutions including
- National Institute of Standards and Technology (NIST)
- LGC Group
- National Measurement Institute (NMI) Australia

• Six tables including a comparison table with specifications of the digital PCR instruments including
- Type of platform
- Number of reactions per sample
- Number of samples the instruments can run or analyze
- Type of detection (end-point or real-time)
- Reaction volume
- Total volume
- Cost per products
- Cost per run
- Cost per sample
- Cost per 10,000 reactions

• Survey results of familiarity of digital PCR and vendors, and expected future use
- Includes up to 220 participants of the following organizations
Academic/University
Biotechnology
Pharmaceutical
Government
Clinical
Other technology providers
- Results depicted over 12 charts and graphs

• Detailed descriptions of the platforms available

• Feedback from end-user communities

Table Of Contents

Table of Contents

Executive Summary

Chapter 1: What is Digital PCR?
1.1 Introduction
1.2 Background Information
1.3 Digital PCR Platforms

Chapter 2: Sensitivity of Digital PCR
2.1 Analyzing Preserved Samples
2.2 Ability to Detect Copy Number Variations (CNVs) and Single Nucleotide Polymorphisms (SNPs)
2.3 Integration with Real-time PCR Assays

Chapter 3: Areas of Improvement for Digital PCR
3.1 Higher Throughput
3.2 Multiplexing
3.3 Recovering Samples
3.4 Cost

Chapter 4: Digital PCR Instruments

Chapter 5: RainDance Technologies
5.1 Background
5.2 RainDrop Digital PCR System
5.3 What is the Process of the RainDrop Digital PCR System?
5.4 Areas of Improvement and Customer Feedback
5.5 Interview With Andy Watson, Chief Commercial Officer at RainDance Technologies
5.5.1 Background
5.5.2 RainDrop Digital PCR System
5.5.3 Sample Preparation and Analysis
5.4.4 Customer Feedback
5.5.5 Future of Digital PCR
5.5.6 Competitive Advantage

Chapter 6: Bio-Rad Laboratories
6.1 Background Information
6.2 QX100 Droplet Digital PCR System
6.3 What is the Process of the QX100 Droplet Digital PCR System?
6.4 Areas of Improvement and Customer Feedback
6.5 Interview With Annette Tumolo, Vice President and General Manager of the Digital Biology Center at Bio-Rad Laboratories
6.5.1 Background
6.5.2 QX100 Dropelet Digital PCR System
6.5.3 Sample Preparation and Analysis
6.5.4 Customer Feedback
6.5.5 Future of Digital PCR
6.5.6 Competitive Advantage

Chapter 7: Life Technologies
7.1 Background
7.2 QuantStudio 3D Digital PCR System
7.3 What is the Process of the QuantStudio 3D Digital PCR System?
7.4 Areas of Improvement and Customer Feedback
7.5 Interview With Mauricio Minotta, Sr. Manager of Corporate Communications at Life Technologies
7.5.1 Background
7.6 Interview With Paco Cifuentes, Director of the Product Application for the Genetic Analysis Business at Life Technologies
7.6.1 QuantStudio 3D Digital PCR System
7.6.2 Sample Preparation and Analysis
7.6.3 Customer Feedback
7.6.4 Future of Digital PCR
7.6.5 Competitive Advantage

Chapter 8: Fluidigm Corporation
8.1 Background
8.2 BioMark HD System vs. EP1 System
8.3 12.765 Digital Array vs. qdPCR 37K IFC
8.4 How do all the Components Work Together?
8.5 What is the Process of the of BioMark HD System?
8.5.1 BioMark HD System
8.5.2 EP1 System
8.6 Areas of Improvement and Customer Feedback
8.7 Interview With Howard High, Senior Fellow, Corporate Communications and Press Officer at Fluidigm Corporation
8.7.1 Background
8.7.2 Digital PCR Systems (BioMark HD and EP1)
8.7.3 Digital PCR Chips (12.765 Digital Array IFC and qdPCR 37K IFC)
8.7.4 Sample Preparation and Analysis
8.7.5 Customer Feedback
8.7.6 Future of Digital PCR
8.7.7 Competitive Advantage

Chapter 9: Comparison of Instrument Specifications

Chapter 10: Design Comparisons

Chapter 11: Digital PCR Use in National Measurement Institutions

Chapter 12: National Institute of Standards and Technology (NIST), United States
12.1 Background
12.2 Poisson Statistics
12.3 Digital PCR Use
12.4 Feedback and Areas of Improvement
12.5 Conclusion
12.6 Interview With Ross Haynes, Technician of Biological Science at National Institute of Standards and Technology (NIST), USA
12.6.1 Background
12.6.2 Poisson Statistics
12.6.3 Digital PCR in NIST Research
12.6.4 Digital PCR Platforms
12.6.5 Digital PCR as a Reference Standard
12.6.6 Digital PCR Pros and Cons

Chapter 13: LGC Group, United Kingdom
13.1 Background
13.2 Advantages
13.3 Precision vs. Accuracy
13.4 Validation of Digital Technology
13.5 Estimation of Molecules
13.6 Sample Collection, Preparation, and Storage
13.7 Inter-Laboratory Comparability
13.8 Feedback and Areas of Improvement
13.9 Conclusion
13.10 Interview With Jim Huggett, Science Leader in Nucleic Acid Metrology at LGC Group, UK
13.10.1 Background
13.10.2 Digital PCR Use
13.10.3 Sample Preparation
13.10.4 Validation
13.10.5 Results
13.10.6 Feedback and Areas of Improvement

Chapter 14: National Measurement Institute (NMI) Australia
14.1 Background
14.2 Reference Standard
14.3 Validation
14.4 Feedback and Areas of Improvement
14.5 Conclusion
14.6 Interview With Kerry Emslie, Manager of the Bioanalysis Group at National Institute of Measurement (NMI) Australia
14.6.1 Background
14.6.2 Digital PCR Application
14.6.3 Validation
14.6.4 Sample Preparation and Analysis
14.6.5 Feedback and Areas of Improvement
14.6.6 Future Outlook

Chapter 15: Digital PCR Use in Research and Development
15.1 Organizations Surveyed
15.2 How well is Digital PCR Known?
15.3 Digital PCR Use in Future Research
15.4 Conclusion
15.5 Survey Questions

References

Acronyms Used In This Report

About Cambridge Healthtech Institute



List of Tables:

Table 3.1: Digital PCR (dPCR) vs. Quantitative real-time PCR (qPCR)
Table 5.1: Summary of RainDrop Digital PCR System
Table 6.1: Summary of QX100 Droplet Digital PCR System
Table 7.1: Summary of QuantStudio 3D Digital PCR System
Table 8.1: Summary of BioMark HD System, EP1 System, 12.765 Digital Array IFC, and qdPCR 37K IFC
Table 9.1: Comparison of Specifications



List of Figures:

Figure 1.1: Flowchart of real-time PCR process
Figure 1.2: Depiction of digital PCR process
Figure 5.1: RainDrop Source: Left. RainDrop Sense: Right
Figure 5.2: RainDrop chip. Up to 8 samples can be loaded
Figure 5.3: Adjusting the concentration of the probes causes them to shift and display different intensities
Figure 5.4: Workflow of RainDrop Digital PCR System
Figure 6.1: QX100 Droplet Digital PCR System components
Figure 6.2: Microfluidics combines oil (North and South channels) with the sample (West channel) to create nanoliter droplets (East channel)
Figure 6.3: Schematic for detecting multiple DNA sequences within a droplet digital PCR fluorescence amplitude scatter plot
Figures 6.4: Positive sequences labeled with FAM (blue), FAM+HEX (green), and HEX (black) dyes
Figure 6.5: Positive sequences labeled with FAM or FAM+HEX (brown), FAM or HEX (blue), and, FAM+HEX or HEX (green) dyes
Figure 6.6: Positive sequences labeled with FAM, FAM+HEX, or HEX dyes
Figure 6.7: Workflow of QX100 Droplet Digital PCR System
Figure 6.8: Step 1: Prepare samples
Figure 6.9: Step 2: Generate droplets using the QX100 droplet generator
Figure 6.10: Step 3: Thermal cycle samples
Figure 6.11: Step 4: Load samples onto the QX100 droplet reader
Figure 6.12: Step 5: Analyze results
Figure 6.13: Step 6: Visualize data
Figure 7.1: QuantStudio 3D Digital PCR System
Figure 7.2: The chip can hold one sample at a time, partitioning it across 20,000 wells for analysis
Figure 7.3: Workflow of QuantStudio 3D Digital PCR System
Figure 8.1: BioMark HD System
Figure 8.2: EP1 System
Figure 8.3: qdPCR 37K IFC
Figure 8.4: Real-time detection captures false positive using the BioMark HD System
Figures 8.5-8.9: Workflow of BioMark HD System

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