SPADA at AOAC Annual Meeting 2023

System Suitability Tests and/or Analytical Quality Control

Sequencing and analysis workflow .— Detection thresholds .—The following thresholds are applied.

AOAC INTERNATIONAL 137 th Annual Meeting & Exposition August 25 – 30, 2023 • New Orleans, LA Criteria for amplicon and organism calls .—To make actionable calls, multiple targets per organism must be detected to achieve strain specificity. For example, Bacillus anthracis requires three targets with all three amplicon reads detected for an actionable call (see Table 18 for required targets per organism). (1) Per alignment threshold: using BLASTN, ≥90% identify across ≥90% alignment length is applied. (2) Per sample (bar code) threshold: a reference must have >2% of total aligned reads; i.e., it is highly unlikely for 50+ amplicons to be truly positive in one sample. (3) Per flow cell threshold: based on NTC aligned read counts per amplicon. Amplicon “detection” will be masked in a sample if sample aligned read count is < (mean NTC + 3 SD) aligned read count.