As of August 15, 2020, PHO Laboratory detected possible false positive SARS-CoV-2 results on
approximately 30 occasions among over 850,000 specimens tested for COVID-19, with ~17,500
(approximately 2%) specimens testing positive. This represents a false positivity rate of less than 0.01%
(specificity of >99.99%), which is well beyond performance targets for a laboratory test, even
acknowledging there are likely to be some false positive tests that go undetected.
In general, the positive predictive value (PPV) of COVID-19 PCR assays is excellent among patients with
high pretest probability, and approaches 100%. This was determined at PHO Laboratory, using viral
sequencing of PCR-positive specimens, excluding those for which viral copy number was near the LOD of
the assay.

An Overview of Cycle Threshold Values
and their Role in SARS-CoV-2 Real-Time PCR Test Interpretation 7

However, in the context of low prevalence when the virus is not circulating at a high level in the
community, PPV drops significantly. For example, if the community prevalence of SARS-CoV-2 is 1%,
with a test sensitivity of 80%, and specificity of 99%, the PPV of a positive test is only 44.7%. If
prevalence increases to 5% or 10%, then the PPV increases significantly to 80.8% and 89.9%,
respectively. A recent serosurvey by PHO using residual convenience specimens found a low adjusted
seroprevalence of 1.1% (95% CI 0.8, 1.3) in June.5 This supports that Ontario is currently a low
prevalence setting for SARS-CoV-2 infection.
False positive results can occur at various stages of laboratory testing, and can be grouped into the
following categories:
1. Pre-analytical errors. These are errors that occur prior to the actual testing and include issues
such as mislabelling of specimens, resulting in incorrect results being reported. These errors may
also result from specimen contamination at time of collection, in transport, or during aliquoting
in the laboratory.
2. Analytical errors. These occur during the laboratory testing process. These include, for
example, errors related to reagent contamination. Reagents can arrive contaminated from the
supplier. To mitigate this, each new lot of reagents undergoes a quality assurance check before
being put into use. False positive results can also arise from contamination due to pipetting
errors, which can be due to human error or defects in automated equipment. Such analytic
errors are controlled through the use of negative and positive controls on each run of the assay,
which are reviewed prior to releasing results.
3. Post-analytical errors. These errors occur at the stage of test result interpretation and
reporting of results. Incorrect interpretation of amplification curves could lead to a false positive
result. Transcription errors may also result in false positive results being generated. Such errors
are mitigated by ensuring all results and interpretations are reviewed prior to reporting.