NAAT on RTB.

The COVID-19 ID Now assay (Abbott Diagnostics, Scarborough, MA) was performed on-site following manufacturer instructions for swab-based collections, except for two drops of RTB that were added to the sample chamber before processing the original Ag-RDT collection swab (Fig. 1). The remaining RTB and swab were transported to a central laboratory in the Ag-RDT reaction tube, and 200 μL of viral transport medial (VTM) (Rodoxica, Little Rock, AR) was added to the tube (to ensure sufficient volumes for NAAT testing). Following vortexing for 10 s, 200 μL of VTM/RTB fluid was subjected to a total nucleic acid extraction (TNA) on a MagNA Pure 96 or LC 2.0 instrument (Roche Diagnostics ltd., Roltkreuz, Switzerland), and 5 μL of the 50 μL of eluted TNAs were used as the template for real-time RT-PCR using the TaqPath COVID-19 Combo kit (Life Technologies Corp., Frederick, MD).