Investigation of pooling strategies using clinical COVID-19 samples for more efficient diagnostic testing

Adikari, S. H.; Alipio Lyon, E. Z.; Hollander, A. D.; Deshpande, A.; Hong-Geller, E.

When testing large numbers of clinical COVID-19 samples for diagnostic purposes, pooling samples together for processing can offer significant reductions in the materials, reagents, time, and labor needed. We have evaluated two different strategies for pooling independent nasopharyngeal swab samples prior to testing with an EUA-approved SARS-CoV-2 RT-qPCR diagnostic assay. First, in the Dilution Study, we assessed the assay's ability to detect a single positive clinical sample diluted in multiple negative samples before the viral RNA extraction stage. We observed that positive samples with Ct values at ~30 can be reliably detected in pools of up to 30 independent samples, and positive samples with Ct values at ~35 can be detected in pools of 5 samples. Second, in the Reloading Study, we assessed the efficacy of reloading QIAamp viral RNA extraction columns numerous times using a single positive sample and multiple negative samples. We determined that one RNA extraction column can be reloaded with up to 20 clinical samples (1 positive and 19 negatives) sequentially without any loss of signal in the diagnostic assay. Furthermore, we found there was no significant difference in assay readout whether the positive sample was loaded first or last in a series of 20 samples. These results demonstrate that different pooling strategies can lead to increased process efficiencies for COVID-19 clinical diagnostic testing.