Author: Andrew C Nelson; Benjamin Auch; Matthew Schomaker; Daryl M Gohl; Patrick Grady; Darrell Johnson; Robyn Kincaid; Kylene E Karnuth; Jerry Daniel; Jessica K Fiege; Elizabeth J Fay; Tyler Bold; Ryan A Langlois; Kenneth B Beckman; Sophia Yohe
Title: Analytical Validation of a COVID-19 qRT-PCR Detection Assay Using a 384-well Format and Three Extraction Methods Document date: 2020_4_5
ID: ec3egn8o_22
Snippet: Quality review of the amplification curves throughout the preliminary assay characterization experiments was performed to optimize the ΔRn threshold value. We noted that artifactual priming events in early amplification cycles would occasionally produce non-exponential amplification traces (Figure 1) . A subset of these non-specific traces could cross if autothresholding set by the instrument was low (ΔRn threshold 0.2-0.3). We noted that all p.....
Document: Quality review of the amplification curves throughout the preliminary assay characterization experiments was performed to optimize the ΔRn threshold value. We noted that artifactual priming events in early amplification cycles would occasionally produce non-exponential amplification traces (Figure 1) . A subset of these non-specific traces could cross if autothresholding set by the instrument was low (ΔRn threshold 0.2-0.3). We noted that all preliminary true positive data points, including the initial limit of detection experiment, demonstrated robust exponential amplification curves which crossed a 0.5 ΔRn threshold prior to 40 cycles; no artifactual traces reached this cutoff. Therefore we set this threshold for all downstream validation experiments.
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