Evaluation of the ddPCR ability to detect the SARS-CoV-2 in nasopharyngeal samples of symptomatic patients with suspected COVID-19 infection using an IgG serological assay (EUROIMMUN Anti-SARS-Cov2 ELISA Ig) as gold/reference standard (FDA validated commercial serologic test).
The Bio-Rad SARS-CoV-2 ddPCR Test is a reverse transcription (RT) droplet digital polymerase
chain reaction (ddPCR) test designed to detect RNA from SARS-CoV-2 in specimens (mainly
nasopharyngeal, anterior nasal, oropharyngeal and mid-turbinate swab but also nasopharyngeal
wash/aspirate and nasal aspirate specimens) collected from individuals who are suspected of
COVID-19 infection.
This single assay multiplex test enables a one-well reaction with three sets of the
oligonucleotide primers and probes which were reported by CDC. Two were selected from regions
of the virus nucleocapsid (N) gene. An additional primer/probe included in the panel is set
to detect the human RNase P gene (RP) in control samples and clinical specimens.
RNA isolated and purified from swab specimens is added to the mastermix comprised of reverse
transcriptase whereby RNA is converted into cDNA and then amplified, using the Bio-Rad
One-Step RT-ddPCR Advanced Kit for Probes.
Briefly, the sample and mastermix RT-ddPCR mixtures are fractionated into up to 20,000
nanoliter-sized droplets in the form of a water-in-oil emulsion. The 96-well RT-ddPCR ready
plate containing droplets is sealed with foil using a plate sealer. The emulsions are then
thermocycled to achieve reverse transcription to generate cDNA followed by target
amplification plus probe hydrolysis in each droplet. After thermocycling is complete, the
96-well RT-ddPCR ready plate is loaded into the Droplet Reader. The Droplet Reader singulates
the droplets and flows them past a two-color fluorescence detector (FAM and HEX) in order to
determine which contain target (positive) and which do not (negative) for each of the targets
identified with the SARS-CoV-2 ddPCR Test: N1, N2 and RP. The ddPCR system uses the
QuantaSoft 1.7 and QuantaSoft Analysis Pro 1.0 for analysis software.
Diagnostic Test: Nasopharyngeal and throat/oropharyngeal swabs analyses by RT-PCR and ddPCR
Nasopharyngeal and throat/oropharyngeal swabs analyzed by both RT-PCR and ddPCR
1. Age ≥ 18 years on the day of signing informed consent.
2. Confirmed diagnosis of any type of solid or hematologic tumor.
3. Ongoing anticancer treatment at the time of inclusion or within the last 3 months
prior to inclusion (last treatment administration or last loco regional procedure).
4. Suspicion of COVID-19 infection. Patients must not have underwent diagnostic test
and/or chest imaging before inclusion.
5. Covered by a medical/health insurance.
6. Signed and dated IRB/ICE approved informed consent form.
Centre Leon Berard
Lyon, France
Bénédicte MASTROIANNI, MD, Principal Investigator
Centre Léon Berard