Stoichiometric Encoding of Amplification Kinetics Enables High-Level Multiplexing in Single-Channel TaqMan Real-Time PCR

Stoichiometric encoding of amplification kinetics enables single-channel 8-plex TaqMan real-time PCR, improving mean silhouette score from 0.329 to 0.711 on a CNS pathogen panel.

• Why it matters: Spectral overlap limits conventional multiplex qPCR, and amplification-curve analysis has been constrained by instrument/specimen domain shifts, limiting reliable multi-target discrimination.
• What they did: They co-designed primer/probe stoichiometry to generate 27-condition reproducible non-canonical amplification curves, then decoded them using a transformer pretrained on 170,000+ unlabeled dqPCR curves with conditional domain adversarial alignment.
• The result: On contrived clinical specimens, classification reached 83.6% accuracy for a canonical 7-plex respiratory virus panel and 89.8% for the encoded 8-plex panel, and co-amplification events were resolved at 98.7% accuracy in controlled dqPCR.

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