Raindance PCR



Built on RainDance’s patented and proven RainStorm™ digital droplet technology,the RainDrop™ System generates up to 10 million picoliter-sized droplets per lane. Since each droplet encapsulates a single molecule, researchers can quickly determine the absolute number of droplets containing specific target DNA and compare that to the number of droplets with background wild-type DNA. The RainDrop System also shifts the current digital PCR (dPCR) paradigm from a single-color-per-marker approach to a two color with varying probe intensity method that is capable of multiplexing up to 10 markers. br/>

Advantages


  • Sensitivity: Detect 1 mutant amongst 250,000 wild-type molecules with a lower limit of detection of 1 in more than 1,000,000.
  • Multiplexing: Conduct up to 10 tests or more on the same sample using the single molecule multi-color detection technique.
  • Study design flexibility: Optimize number of PCR reactions based on your sensitivity AND multiplex requirements.
  • Closed-tube design: Ensure highest quality by eliminating contamination or carryover.
  • Proven technology platform: Leverage a decade of experience with the same core technology found in RainDance’s Targeted Sequencing Systems.
  • Cost per data point: Generate true digital answers with orders of magnitude more data per dollar.

RainDance Droplet PCR Workflow



Capable of generating more than a billion reactions in a single day, the RainDrop® Digital PCR System surpasses all existing technologies and establishes a new performance standard in sensitivity, quantitation and multiplexing. Based on RainDance’s proven digital droplet platform, the RainDrop System fundamentally changes the performance of molecular assays by enabling digital answers across a number of important applications including low-frequency tumor allele detection,gene expression, copy number variation, and SNP measurement.

Innovative Design

A new gold standard in sensitivity is only possible through the careful consideration of every aspect of the system. The RainDrop System offers a number of innovative digital droplet-based design advantages.

RainDrop

  • Automated delivery of filtered oil from sealed on-board reservoirs eliminates any contamination risks from the environment or pipetting
  • Real-time closed-loop control of droplet creation ensures uniformity both within and across runs
  • Automated filling of PCR tubes with emulsion avoids possible losses with manual pipetting techniques
  • All wetted surfaces are single use, eliminating the potential for any carryover of droplets from one sample to the next that can occur with reusable fluid path approaches
  • Sealed amplicon containment completely eliminates catastrophic contamination issues from spills and other problems with pierced foil designs
  • Logical channel-to-channel workflow, integrated bar-coding and sample tracking through the RainDrop software eliminate the risk of experiment disasters caused by simple reformatting mistakes in sample handling.

Robust and Reliable Platform

Don’t risk your research experiments to first-generation designs. The RainDrop System is the third generation of RainDance’s digital droplet-based PCR instrumentation and uses the fourth generation of our microfluidic chips. These innovations have a long track record of success in the lab and have been detailed by leading research institutions in publications including Analytical Chemistry, Lab on a Chip, and PNAS.


Streamlined Workflow

RainDrop Digital PCR Workflow

The RainDrop System shifts the PCR paradigm from a single color per marker approach to a more scalable and precise multi-color and intensity-per-marker method. This novel approach increases sensitivity by generating between 1 million and 10 million pico-liter sized droplets per lane, which is a 500 – 10,000x improvement over existing PCR methods. Since each droplet encapsulates a single molecule, researchers can quickly determine the absolute number of droplets containing specific target DNA and compare that to the amount of droplets with normal, background wild-type DNA.


Sensitivity

To assess the lower limit of detection (LLOD) of an EGFR mutation, varying concentrations of mutant were diluted into genomic wild-type DNA. The assay delivered a linear response (R2 = 0.998) down to 1 mutant amongst 250,000 wild-type molecules, with a LLOD of 1 in more than 1,000,000 as defined by the average of the wild-type only controls plus 3x the standard deviation.


Multiplexing

By adjusting concentrations of the probes for each individual assay, 4 mutations plus wild-type were measured simultaneously with just VIC and FAM fluorophores. Multiplexing can be expanded to higher plex levels, and it enables detection, identification, and measurement of multiple possible mutations from a single DNA sample.


Video of Droplet Creation on RainDrop Source

  • 80,000 PCR reactions per second – Droplets are created in all 8 channels simultaneously at rates approaching 5 million per minute.
  • CCD vision system provides real-time closed-loop control to ensure droplet size uniformity.
  • Droplet data retained in Source log files for traceability.

The RainDrop Digital PCR System is for Research Use Only; not for use in diagnostic procedures.

This application note, Human Universal RNA, Xeno RNA and de-identified samples from some of our key customers were evaluated to demonstrate the RainDrop’s utility for sensitive, precise multiplex detection of HIV DNA or RNA.


Multiplexing
Multiplexing with RainDrop Digital PCR
The RainDrop™ Digital PCR System provides unique capabilities and benefits for counting applications such as rare mutation detection, quantifying copy number variation, DNA methylation assessment, and as a pre-test QC and validation tool for DNA sequencing. The power of digital PCR is extended even further with the novel multiplexing capability made possible with millions of droplets. This application note summarizes the simple approach used to develop multiplex assays and provides examples of important multiplexed assays in various research applications.



MicroRNA
Quantifying MicroRNA Using RainDrop™ Digital PCR System
Unlike most other potential biomarkers, microRNA molecules appear to be cell type and disease specific, as well as stable and readily accessible. This application note demonstrates the use of RainDrop™ Digital PCR System as an exciting new tool for microRNA research.


Validation Tool for Sequencing
RainDrop™ Digital PCR System as a Pre-test QC & Validation Tool for Sequencing
This application note describes two droplet-based digital PCR methods which provide significant value for DNA sequencing. First, a dPCR DNA Quality Assay is described that can be used for simultaneous detection of DNA integrity and the quantity of amplifiable DNA, enabling cost-effective sequencer use. Second, digital PCR provides a simple and sensitive orthogonal method for validation of sequencing results that show low frequency minor alleles.


Copy Number Variation
Digital PCR Multiplexing Assay for Gene Copy Number Research
This application note presents the results of a novel 5-plex droplet-based digital PCR assay to identify gene copy variations and SNPs. Assay performance demonstrates accurate quantification and superior reproducibility across a set of samples and a wide range of copy number levels.



Biomarkers
RainDrop Digital PCR System Quantifies Methylated DNA Biomarkers
This application note illustrates the RainDrop™ Digital PCR System’s capability to quantify small changes in DNA methylation. Highlighted is analysis using Zymo Research’s OneStep qMethyl™ kit with low amounts of tumor DNA in single- and multiplex assay formats.


VIC
Using Alternatives to VIC on the RainDrop™ Digital PCR System
The RainDrop™ Digital PCR System allows users to pair a second probe in conjunction with FAM. While VIC is the most commonly paired second probe, this application note highlights 6 VIC alternatives and their intensity relative to VIC on the RainDrop Digital PCR System.


OneStep
Using the RainDrop™ Digital PCR System for One-Step RT-dPCR
One-step RT-dPCR has many advantages over a two-step reverse transcriptase protocol – namely: 1) simpler workflow; 2) reduced variability due to fewer pipetting and purification steps; 3) reduces 3’ and 5’ biases introduced by random oligomers and oligo-dT primers; 4) single RNA molecules converted to cDNA without competition. The RainDrop™ Digital PCR System provides absolute quantification, removing the need for a standard curve, and has a broad dynamic range that allows for quantification of both high expressing and low expressing genes in a single well. In this study, four commercially available RT-PCR kits were tested in the RainDrop Digital PCR System: SuperScript® III One-Step RT-PCR System (Life Technologies); AgPath-ID™ One-Step RT-PCR Kit (Life Technologies); qScriptTM XLT One-Step RT-qPCR Toughmix® (Quanta BioSciences); and QIAGEN® OneStep RT-PCR Kit (Qiagen). While all kits performed well, this application note presents experimental data for the SuperScript® III One-Step RT-PCR Kit using its specified protocol only.


HpCNV
High-Precision Chromosomal Copy Number Measurement Research using RainDrop™ Digital PCR System
This application note discusses using the RainDrop™ Digital PCR System to overcome these measurement barriers and precisely measure chromosomal copy number differences. The most extreme case demonstrated is the discernment of affected and unaffected samples with a 99% confidence interval when the affected fraction of a sample makes up <5% of the 10ng starting material.


RainDance RainDrop System consumables are designed and manufactured to the highest standards to provide leading-edge performance and reliability.

Source Chip

The Source Chip: Quality begins at the source

  • Droplet generation rate approaches 5 million per minute (8 samples)
  • Parallel design of flow in channels enables imaging via CCD camera for feedback and precision control
  • Sealing caps are installed in a clean environment to eliminate contamination risk

Sense Chip: Repeatable identification and counting of droplets

    Sense Chip
  • Submicron tolerance nozzles automatically align and space droplets for robust operation
  • Auto-piercing of the caps and closed tube design seals the amplicons, preventing any contamination of the environment or other samples
  • COC polymer provides superior signal to noise detection

Proprietary Oil Surfactants – Increase stability and performance

  • Proprietary surfactants enable 5 pL droplets with superior stability and minimal coalescence under PCR conditions
  • Proven across multiple commercial products
  • Rigorous QC and extended shelf life
  • Novel syringe packaging makes for easy refilling of RainDrop reservoirs

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