How to multiplex with color combination in Crystal Digital PCR®?

In color combination multiplexing, a target is identified using two separate fluorophores detected in 2 different channels. By using two probes per sequence of interest, either competing or
non-competing, each population can be revealed in 2 colors, expanding the number of targets per assay detectable beyond the
number of available channels (Figure 1A).

With Nio® Digital PCR, up to 21 targets can be detected with its 7-color channels.

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Technical Note Highlights

• Color combination allows to detect and quantify up to 21 targets in a single reaction with Nio® Digital PCR.
• Nio® Analyzer Software unique feature, population editor, allows for single thresholding between positive and negative populations.
• Tips for assay design in color combination: application-specific design strategy, smart assay configuration, and low co-encapsulation rate.
• Color combination approach allows for the development of hyperplexed assays, such as the 18-plex ESR1 (17 mutations) Crystal Digital PCR® Assay.