Dried blood spot specimens for SARS-CoV-2 antibody testing: A multi-site, multi-assay comparison

Abstract

The true severity of infection due to COVID-19 is under-represented because it is based on only those who are tested. Although nucleic acid amplifications tests (NAAT) are the gold standard for COVID-19 diagnostic testing, serological assays provide better population-level SARS-CoV-2 prevalence estimates. Implementing large sero-surveys present several logistical challenges within Canada due its unique geography including rural and remote communities. Dried blood spot (DBS) sampling is a practical solution but comparative performance data on SARS-CoV-2 serological tests using DBS is currently lacking. Here we present test performance data from a well-characterized SARS-CoV-2 DBS panel sent to laboratories across Canada representing 10 commercial and 2 in-house developed tests for SARS-CoV-2 antibodies. Three commercial assays identified all positive and negative DBS correctly corresponding to a sensitivity, specificity, positive predictive value, and negative predictive value of 100% (95% CI = 72.2, 100). Two in-house assays also performed equally well. In contrast, several commercial assays could not achieve a sensitivity greater than 40% or a negative predictive value greater than 60%. Our findings represent the foundation for future validation studies on DBS specimens that will play a central role in strengthening Canada's public health policy in response to COVID-19.

Fig 1. Positive predictive values (PPV) and negative predictive values (NPV) by prevalence for each commercial and in-house assay on dried blood spot specimens.

PPV and NPV is presented for n = 10 SARS-CoV-2 antibody negative DBS specimens and n = 10 SARS-CoV-2 antibody positive DBS specimens. One 6 mm (1/4 inch) punch was used for the EUROIMMUN assay and two 6 mm (1/4 inch) punches were used for the Platelia and in-house assays. Data from four 6 mm (1/4 inch) punches is presented for all other assays. EUROIMMUN = Anti-SARS-CoV-2 ELISA assay (EUROIMMUN, Lübeck, Germany). Platelia = SARS-CoV-2 assay (Bio-Rad, Hercules, California). LIASON = SARS-CoV-2 assay (DiaSorin, Saluggia, Italy). COV2G = SARS-CoV-2 COV2G assay (Siemens, Erlangen, Germany). COV2T = SARS-CoV-2 COV2T assay (Siemens). Elecsys S = Quantitative Anti-SARS-CoV-2 assay (Elecsys spike; Roche, Basel, Switzerland). Elecsys N = Anti-SARS-CoV-2 assay (Elecsys nucleocapsid; Roche). VITROS = Anti-SARS-CoV-2 assay (Ortho Clinical Diagnostics, Raritan, New Jersey). Architect = SARS-CoV-2 assay (Abbott, Mississauga, Canada). GSP/DELFIA = Anti-SARS-CoV-2 assay (PerkinElmer, Waltham, Massachusetts). In-house S (U of T) = In-house spike assay (University of Toronto). In-house RBD (U of T) = In-house RBD assay (University of Toronto). In-house N (U of T) = In-house nucleocapsid assay (University of Toronto). In-house S, mono (U of O) = In-house monoclonal spike assay (University of Ottawa). In-house RBD, mono (U of O) = In-house monoclonal RBD assay (University of Ottawa). In-house N, mono (U of O) = In-house monoclonal nucleocapsid assay (University of Ottawa). The VITROS assay could not achieve a specificity greater than 0% therefore, only PPV is shown.

Fig 2

Inter-assay concordance (A) and Cohen’s kappa coefficient (B) for SARS-CoV-2 antibody testing on dried blood spot specimens. Concordance (%) and Cohen’s kappa coefficient are presented for n = 10 SARS-CoV-2 antibody negative DBS specimens and n = 10 SARS-CoV-2 antibody positive DBS specimens. One 6 mm (1/4 inch) punch was used for the EUROIMMUN assay and two 6 mm (1/4 inch) punches were used for the Platelia and in-house assays. EUROIMMUN = Anti-SARS-CoV-2 ELISA assay (EUROIMMUN, Lübeck, Germany). Platelia = SARS-CoV-2 assay (Bio-Rad, Hercules, California). LIASON = SARS-CoV-2 assay (DiaSorin, Saluggia, Italy). COV2G = SARS-CoV-2 COV2G assay (Siemens, Erlangen, Germany). COV2T = SARS-CoV-2 COV2T assay (Siemens). Elecsys S = Quantitative Anti-SARS-CoV-2 assay (Elecsys spike; Roche, Basel, Switzerland). Elecsys N = Anti-SARS-CoV-2 assay (Elecsys nucleocapsid; Roche). VITROS = Anti-SARS-CoV-2 assay (Ortho Clinical Diagnostics, Raritan, New Jersey). Architect = SARS-CoV-2 assay (Abbott, Mississauga, Canada). GSP/DELFIA = Anti-SARS-CoV-2 assay (PerkinElmer, Waltham, Massachusetts). In-house S (U of T) = In-house spike assay (University of Toronto). In-house RBD (U of T) = In-house RBD assay (University of Toronto). In-house N (U of T) = In-house nucleocapsid assay (University of Toronto). In-house S, mono (U of O) = In- house monoclonal spike assay (University of Ottawa). In-house RBD, mono (U of O) = In-house monoclonal RBD assay (University of Ottawa). In-house N, mono (U of O) = In-house monoclonal nucleocapsid assay (University of Ottawa).

Fig 3. Receiver operating characteristic (ROC) curve for each commercial and in-house assay on dried blood spot specimens.

ROC curves are presented for n = 10 SARS-CoV-2 antibody negative DBS specimens and n = 10 SARS-CoV-2 antibody positive DBS specimens. One 6 mm (1/4 inch) punch was used for the EUROIMMUN assay and two 6 mm (1/4 inch) punches were used for the Platelia and in-house assays. Data from four 6 mm (1/4 inch) punches is presented for all other assays. EUROIMMUN = Anti-SARS-CoV-2 ELISA assay (EUROIMMUN, Lübeck, Germany). Platelia = SARS-CoV-2 assay (Bio-Rad, Hercules, California). LIASON = SARS-CoV-2 assay (DiaSorin, Saluggia, Italy). COV2G = SARS-CoV-2 COV2G assay (Siemens, Erlangen, Germany). COV2T = SARS-CoV-2 COV2T assay (Siemens). Elecsys S = Quantitative Anti-SARS-CoV-2 assay (Elecsys spike; Roche, Basel, Switzerland). Elecsys N = Anti-SARS-CoV-2 assay (Elecsys nucleocapsid; Roche). VITROS = Anti-SARS-CoV-2 assay (Ortho Clinical Diagnostics, Raritan, New Jersey). Architect = SARS-CoV-2 assay (Abbott, Mississauga, Canada). GSP/DELFIA = Anti-SARS-CoV-2 assay (PerkinElmer, Waltham, Massachusetts). In-house S (U of T) = In-house spike assay (University of Toronto). In-house RBD (U of T) = In-house RBD assay (University of Toronto). In-house N (U of T) = In-house nucleocapsid assay (University of Toronto). In-house S, mono (U of O) = In-house monoclonal spike assay (University of Ottawa). In-house RBD, mono (U of O) = In-house monoclonal RBD assay (University of Ottawa). In-house N, mono (U of O) = In-house monoclonal nucleocapsid assay (University of Ottawa).

Fig 4. Distribution of values obtained for each commercial and in-house assay on dried blood spot (DBS) specimens.

Each panel shows the optical density ratio (OD Ratio), arbitrary units per mL (AU/mL), index, units per mL (U/mL), cut-off index, or signal to cut-off ratio (S/Co) for SARS-CoV-2 antibody negative DBS specimens (n = 10) represented in blue and SARS-CoV-2 antibody positive DBS specimens (n = 10) represented in orange. All values are log₁₀ transformed to aid with visualisation. One 6 mm (1/4 inch) punch was used for the EUROIMMUN assay and two 6 mm (1/4 inch) punches were used for the Platelia and in-house assays. Data from four 6 mm (1/4 inch) punches is presented for all other assays. EUROIMMUN = Anti-SARS-CoV-2 ELISA assay (EUROIMMUN, Lübeck, Germany). Platelia = SARS-CoV-2 assay (Bio-Rad, Hercules, California). LIASON = SARS-CoV-2 assay (DiaSorin, Saluggia, Italy). COV2G = SARS-CoV-2 COV2G assay (Siemens, Erlangen, Germany). COV2T = SARS-CoV-2 COV2T assay (Siemens). Elecsys S = Quantitative Anti-SARS-CoV-2 assay (Elecsys spike; Roche, Basel, Switzerland). Elecsys N = Anti-SARS-CoV-2 assay (Elecsys nucleocapsid; Roche). VITROS = Anti-SARS-CoV-2 assay (Ortho Clinical Diagnostics, Raritan, New Jersey). Architect = SARS-CoV-2 assay (Abbott, Mississauga, Canada). GSP/DELFIA = Anti-SARS-CoV-2 assay (PerkinElmer, Waltham, Massachusetts). In-house S (U of T) = In-house spike assay (University of Toronto). In-house RBD (U of T) = In-house RBD assay (University of Toronto). In-house N (U of T) = In-house nucleocapsid assay (University of Toronto). In-house S, mono (U of O) = In-house monoclonal spike assay (University of Ottawa). In-house RBD, mono (U of O) = In-house monoclonal RBD assay (University of Ottawa). In-house N, mono (U of O) = In-house monoclonal nucleocapsid assay (University of Ottawa).