Achieve Reliable Precision in ddPCR Workflows

Bulletin 3530 Thermal Cycling Abstract This study demonstrates the thermal performance between the PTC Tempo Deepwell Thermal Cycler and the C1000 Touch Thermal Cycler with 96–Deep Well Reaction Module (C1000 Touch 96–Deep Well Thermal Cycler) for amplification of input DNA prior to Droplet Digital PCR (ddPCR™) applications on the QX200™ and QX600™ Droplet Digital PCR Systems. The Bio-Rad QX600 ddPCR System delivers best-in-class performance with the addition of six-color multiplexing. A previous study determined thermal efficiency across a gradient, ramp rate, accuracy, and gradient performance, and demonstrated comparable thermal performance of the PTC Tempo Deepwell and C1000 Touch 96–Deep Well Thermal Cyclers (bulletin 3483). In this evaluation, the downstream amplification products provided by the PTC Tempo Deepwell and C1000 Touch 96–Deep Well Thermal Cyclers were measured for their use in a ddPCR workflow performed on the QX200 and QX600 ddPCR Systems, to determine the resulting accuracy and precision of the two formats. This comparability study demonstrates that the accuracy and precision of measurement of the DNA amplification products generated are comparable between the PTC Tempo Deepwell and C1000 Touch 96–Deep Well Thermal Cyclers. The PTC Tempo Deepwell Thermal Cycler is validated to support the Bio-Rad ddPCR workflow, specifically the QX200 and QX600 Droplet Digital PCR Systems. Introduction For this evaluation of compatibility with Droplet Digital PCR, the PTC Tempo Deepwell Thermal Cycler is used to amplify the DNA target to endpoint within each droplet partition. After droplet generation and PCR, each partition is read using a droplet reader (either with the QX200 or QX600 Droplet Reader) to determine the fraction of positive partitions that includes the target for calculation of target DNA concentration. To ensure robust amplification, PTC Tempo Thermal Cyclers were designed to include precise temperature control in a flexible thermal gradient configuration. For DNA amplification in Droplet Digital PCR, the PTC Tempo Deepwell Thermal Cycler is preferred as it accommodates larger volumes and provides a greater amplification yield. Specifically, the deepwell reaction module offers a block height that ensures homogenous heating of the droplet emulsion during Droplet Digital PCR. The PTC Tempo Deepwell Thermal Cycler builds on the performance of its predecessor, the C1000 Touch 96–Deep Well Thermal Cycler, and now includes user-friendly features such as an automatic lid capable of autosensing plates and tubes, an LED light display, audible status notifications, and connectivity options that include the Bio-Rad BR.io cloud platform, WiFi, USB, and Ethernet connections. Moreover, the PTC Tempo Deepwell Thermal Cycler maintains the same user access controls for user management with the addition of enhanced security and reporting features. This study evaluates the consistency in DNA concentration measurements of samples obtained on the QX200 and QX600 ddPCR Systems using the PTC Tempo Deepwell Thermal Cycler and the C1000 Touch 96–Deep Well Thermal Cycler. The study shows its ability to provide high accuracy and precision of a DNA target in ddPCR applications. A Comparability Study of the C1000 Touch 96–Deep Well and PTC Tempo Deepwell Thermal Cyclers for Use in Droplet Digital™ PCR Workflow