For applications that require a high degree of precision, sensitivity, and reproducibility, The CONSTELLATION® Digital PCR system offers a user-friendly, high-throughput, and speedy platform for the absolute quantification of up to five nucleic acid targets. However, in order for the platform to be useful, the dynamic range of the instrument must exceed the range of sample concentrations to be analyzed for a given assay. In order to provide researchers with the flexibility to trade-off dynamic range for sample throughput, two different CONSTELALTION Digital PCR plates can be run on the platform.
This applications note highlights the dynamic range of the Standard Digital PCR plates as a guide to help researchers understand whether the Standard Digital PCR plate offers sufficient dynamic range for their application. Researchers may utilize High-Dynamic Range plates for applications such as rare-target detection that require quantification of low abundance targets in a background a highly abundant housekeeping gene.
As evidenced by an R2 value of .9921, the combination of reproducible liquid transfers and precise digital quantification approaches provide reliable data across a wide dynamic range from an initial sample concentration of 10 to over 50,000 copies of target DNA per microliter. Although the precision of quantification deteriorates at low concentrations of target DNA, the CONSTELLATION Standard Digital PCR plates offer precision within 10% in all final reaction concentrations above 40 copies per microliter, which corresponds to a starting sample concentration of 100 copies per microliter. For all concentrations below this, the CONSTELLATION High-dynamic Range Plates are recommended.
- From 40 copies/microliter to 23,500 copies/microliter, the precision of measurement of the CONSTELLATION is within 10%.
- The dynamic range assessed in this study showcases the CONSTELLATION as a useful quantification platform for gene expression analysis, NGS Library Quantification, Low Copy Number Variation, and Monitoring CRISPR Gene Editing Projects.
- Sample preparation for the CONSTELLATION mirrors that of traditional qPCR, and can be efficiently and reproducibly automated using the Gilson PIPETMAX and MANTIS Liquid Handlers.
- Dead volume of master mix was only 6 µL due to the novel positive displacement technology employed by the MANTIS Liquid Handler.
- Automated sample dilutions and plate loading on PIPETMAX® ensure accurate and consistent sample and reaction plate pipetting.
- The Luna Universal Probe qPCR Master Mix format simplifies reaction setup and offers exceptional sensitivity and reproducibility.