Lumipulse G SARS-CoV-2 Ag assay evaluation using clinical samples from different testing groups

  1. Giulia Menchinelli
  2. Licia Bordi
  3. Flora Marzia Liotti
  4. Ivana Palucci
  5. Maria Rosaria Capobianchi
  6. Giuseppe Sberna
  7. Eleonora Lalle
  8. Lucio Romano
  9. Giulia De Angelis
  10. Simona Marchetti
  11. Maurizio Sanguinetti
  12. Paola Cattani
  13. Brunella Posteraro

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Abstract

Objectives

Compared to RT-PCR, lower performance of antigen detection assays, including the Lumipulse G SARS-CoV-2 Ag assay, may depend on specific testing scenarios.

Methods

We tested 594 nasopharyngeal swab samples from individuals with COVID-19 (RT-PCR cycle threshold [Ct] values ≤ 40) or non-COVID-19 (Ct values >40) diagnoses. RT-PCR positive samples were assigned to diagnostic, screening, or monitoring groups of testing.

Results

With a limit of detection of 1.2 × 10 4 SARS-CoV-2 RNA copies/mL, Lumipulse showed positive percent agreement (PPA) of 79.9% (155/194) and negative percent agreement of 99.3% (397/400), whereas PPAs were 100% for samples with Ct values of <18 or 18–<25 and 92.5% for samples with Ct values of 25–<30. By three groups, Lumipulse showed PPA of 87.0% (60/69), 81.1% (43/53), or 72.2% (52/72), respectively, whereas PPA was 100% for samples with Ct values of <18 or 18–<25, and was 94.4, 80.0, or 100% for samples with Ct values of 25–<30, respectively. Additional testing of RT-PCR positive samples for SARS-CoV-2 subgenomic RNA showed that, by three groups, PPA was 63.8% (44/69), 62.3% (33/53), or 33.3% (24/72), respectively. PPAs dropped to 55.6, 20.0, or 41.7% for samples with Ct values of 25–<30, respectively. All 101 samples with a subgenomic RNA positive result had a Lumipulse assay’s antigen positive result, whereas only 54 (58.1%) of remaining 93 samples had a Lumipulse assay’s antigen positive result.

Conclusions

Lumipulse assay was highly sensitive in samples with low RT-PCR Ct values, implying repeated testing to reduce consequences of false-negative results.

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  1. Version published to 10.1515/cclm-2021-0182
  2. ScreenIT

    SciScore for 10.1101/2021.01.26.21250533: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board StatementIRB: Study design and clinical samples: This study was conducted at the Fondazione Policlinico Universitario A. Gemelli IRCCS (FPG) and was approved by the FPG Ethics Committee (reference number 49978/20).
    Consent: Informed consent was obtained from all participants before including their samples in the study.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    In the first reaction, the sample (or the SARS-CoV-2 Ag calibrator) and the sample treatment solution are added to an anti-SARS-CoV-2 monoclonal antibody-coated magnetic particle solution, and then incubated for 10 min at 37°C to allow formation of specific antigen-antibody immunocomplexes.
    anti-SARS-CoV-2 monoclonal antibody-coated magnetic particle solution,
    suggested: None
    In the second reaction (accessed after washing), an alkaline phosphatase-labelled anti-SARS-CoV-2 monoclonal antibody solution is added and incubated for 10 min at 37°C to allow specific binding to the antigen of aforementioned immunocomplexes, and then to form additional immunocomplexes.
    anti-SARS-CoV-2
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Briefly, aforementioned contrived samples were spiked with a dilution series of Vero E6 cell-cultured SARS-CoV-2 (INMI-1 strain) at a concentration range of 1.0 × 105 50% tissue culture infective dose (TCID50)/ml (4.0 × 108 RNA copies/ml) to 1.0 TCID50/ml (4.0 × 103 RNA copies/ml), and then tested in replicates (Fig. S1).
    Vero E6
    suggested: None
    Software and Algorithms
    SentencesResources
    To determine Lumipulse assay’s LOD, the MedCalc statistical software (MedCalc Software Ltd, Ostend, Belgium) was used to convert RT-PCR positive detection proportion into a “probability unit” (or “probit”).
    MedCalc
    suggested: (MedCalc, RRID:SCR_015044)
    Statistical analysis was conducted using Stata 15 (StataCorp, College Station, TX) or GraphPad Prism 7 (GraphPad Software, San Diego, CA) software. P <0.05 was considered statistically significant.
    StataCorp
    suggested: (Stata, RRID:SCR_012763)
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    GraphPad
    suggested: (GraphPad Prism, RRID:SCR_002798)

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    However, we compensated for this limitation by including testing groups that were comparable for size (∼60 RT-PCR positive samples per group), and we assumed that RT-PCR negative samples were almost equally distributed across testing groups. To summarize, our results show that Lumipulse assay’s performance was satisfactory, confirming the current view about antigen-based laboratory testing for SARS-CoV-2 detection. In particular, the Lumipulse assay was highly sensitive to detect SARS-CoV-2 antigen in samples with low RT-PCR Ct values (<25) by overall or different testing scenarios. While Ct values >25 might not correspond to situations with active SARS-CoV-2 infection and/or infectivity, a strategy of repeated testing can maximize the Lumipulse assay’s performance and thereby reduce consequences of false-negative results.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    About SciScore

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  3. Version published to 10.1101/2021.01.26.21250533 on medRxiv