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Updated: Oct 30, 2025
Large-Scale SARS-CoV-2 Testing Utilizing Saliva and Transposition Sample Pooling
Published on: June 23, 2022
The Hologic Aptima SARS-CoV-2 assay enables high ratio pooling saving reagents and improving turnaround time
Kimberly Newsom1, Yuan Zhang1, Srikar Chamala1
1Department of Pathology, Immunology and Laboratory Medicine, University of Florida, Gainesville, FL, USA.
Background:
The Hologic Aptima™ TMA SARS-CoV-2 assay was employed to test pooled nasopharyngeal (NP) samples to evaluate the performance of pooled sample testing and characterize variables influencing results.
Methods:
Results on 1033 previously tested NP samples were retrieved to characterize the relative light units (RLU) of SARS-CoV-2-positive samples in the tested population. The pooling strategy of combining 10 SARS-CoV-2 samples into one pool (10/1) was used in this study. The results were compared with neat sample testing using the same Aptima™ TMA SARS-CoV-2 assay and also the CDC RT-PCR and the Cepheid SARS-CoV-2 assays.
Results:
The Aptima assay compares favorably with both CDC RT-PCR and the Cepheid SARS-CoV-2 assays. Once samples are pooled 10 to 1 as in our experiments, the resulting signal strength of the assay suffers. A divide opens between pools assembled from strong-positive versus only weak-positive samples. Pools of the former can be reliably detected with positive percent agreement (PPA) of 95.2%, while pools of the latter are frequently misclassified as negative with PPA of 40%. When the weak-positive samples with kRLU value lower than 1012 constitute 3.4% of the total sample profile, the assay PPA approaches 93.4% suggesting that 10/1 pooled sample testing by the Aptima assay is an effective screening tool for SARS-CoV-2.
Conclusion:
Performing pooled testing, one should monitor the weak positives with kRLU lower than 1012 or quantification cycle (Cq) value higher than 35 on an ongoing basis and adjust pooling approaches to avoid reporting false negatives.
