A method comparison study of the high throughput automated HISCL® SARS-CoV-2 antigen assay using nasopharyngeal swab samples from symptomatic and asymptomatic subjects against conventional RT-PCR

Abstract

Our study aim was to evaluate the performance of the automated Sysmex HISCL® severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigen assay against reverse-transcription polymerase chain reaction (RT-PCR). We tested 277 remnant frozen nasopharyngeal swab samples, stored in universal transport medium (UTM), yielding a sensitivity of 94.9% against historical RT-PCR results with cycle threshold (C_t ) < 30, and a sensitivity of 76.7% for C_t < 35, and specificity of 100% (all C_t values) confirming compatibility of UTM-diluted samples with the assay system. Thereafter, we prospectively collected 141 nasopharyngeal swab samples in UTM from healthcare workers and 1369 paired swabs (400 UTM; 969 dry) from individuals at a public health testing center, with the first swab (UTM) reserved for RT-PCR, yielding a positivity rate of 4.6%. HISCL assay performance using UTM swabs was superior to dry swabs, with a sensitivity of 100% (95% confidence interval [CI] 71.5%-100%) at C_t < 30 versus 92.3% (95%CI 81.5%-97.9%), and a specificity of 99.3% (95% CI 98.1-99.89) against 83.3% (95%CI 80.7%-85.6%). We conclude that this antigen assay is suitable for high throughput facilities where the primary indication for testing is to rule out infection with low RT-PCR C_t values (proxy for high viral loads) to curb viral spread.

Keywords: HISCL automated antigen assay; RT-PCR; SARS-CoV-2; method comparison; rapid testing; variants of concern.

Figure 1

Schematic outline of the study sample processing workflows. (A) Part 1—frozen bio‐banked nasopharyngeal swab samples; (B) Part 2a—prospectively collected single nasopharyngeal swabs (UTM) in the hospital setting; (C) Part 2b—prospectively collected double nasopharyngeal swabs (UTM and dry) in the public health testing center. Here a subset of samples underwent filtration only. (D) Part 2c—prospectively collected double nasopharyngeal swabs (UTM only) in the public health testing center. RT‐PCR, reverse transcription‐polymerase chain reaction; UTM, universal transport medium

Figure 2

Comparison of HISCL SARS‐CoV‐2 antigen assay C.O.I values with RT‐PCR C _t values for retrospective testing of bio‐banked nasopharyngeal swabs. The open black dots represent samples with wild‐type SARS‐CoV‐2 infections. The red dots represent samples with the B1.1.177 mutation (n = 22/29) and the blue dots collectively represent samples. C.O.I., cutoff index; RT‐PCR, reverse‐transcription polymerase chain reaction; SARS‐CoV‐2, severe acute respiratory syndrome coronavirus 2

Figure 3

Comparison of HISCL SARS‐CoV‐2 antigen assay C.O.I. values against RT‐PCR C _t values for Part 1, 2a, and 2c and 2b samples combined (n = 1787). The dotted line represents the cutoff between positive (on or above) and negative (below) antigen test results. C.O.I., cutoff index; RT‐PCR, reverse‐transcription polymerase chain reaction; SARS‐CoV‐2, severe acute respiratory syndrome coronavirus 2