Resumen de Evaluation of a total antibody assay against SARS-COV-2
José Luis Martín Calderón, Jesús Timón Zapata, P. Patiño, María Teresa Gil Ruiz
Background-Aim: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a novel coronavirus, emerged in December 2019 in China which is responsible for Coronavirus disease 19 (COVID-19). The virus rapidly spread all over the world and the WHO declared COVID-19 a pandemic in March 2020. Reverse-transcriptase real-time polymerase chain reaction (RTPCR) on respiratory tract samples is the method of choice for detecting SARSCoV-2 infection. However serological test can play an important role in identifying persons with prior SARS-CoV-2 infection, supporting the diagnosis in cases of RT-PCR false negatives and may be used in seroprevalence surveys. In this study we assess an electrochemiluminescent method for measuring total antibodies against SARS-COV-2 nucleocapsid protein, Methods: Determinations were carried-out using an electrochemilumienescent method on the cobas e801 (Roche diagnostics®), which measures total anti-SARS-CoV-2 antibodies against nucleocapsid protein (N), including IgG, IgM and IgA. Antibodies on the sample bind to biotinylated recombinant protein N, labeled with a ruthenium complex.
Electrochemiluminescence is produced when streptavidin-coated particles bind the complex on the electrochemical cell. Results are expressed as a cut-off index (COI), considering positive ≥1 and negative <1. A total of 139 serum samples obtained from patients with RT-PCR performed at least 10 days before were analyzed on the Cobas e801.
Intra and Inter-assay precision was determinate using two samples pools with different antibody title measuring in quintuplicate during five consecutive days according to the Clinical and Laboratory Standards Institute (CLSI) EP15-A3 protocol. Diagnostic sensitivity was performed using the RT-PCR result as a gold standard. In addition we carried-out an interference study by spiking haemolysate, bilirubin and Intralipid 20%, producing an interfering concentration of 1000 mg/dL haemogobin, 30 mg/dL bilirubin and 1625 mg/dl triglycerides. Comparison of samples with and without interfering was performed using the non-parametric Wilcoxon test. Calculations were carried-out using EXCEL 2013 and the statistics package MedCalc 11.3 (Ostend, Belgium) Results: The number of antibodies test positives with prior PCR test positive was 66 (over all 73 positive antibodies test), giving as result diagnostic sensitivity of 90.4%. Intra- and interassay imprecision obtained were respectively 0.57 and 7.01 (low title pool), and 2.79 and 5.11 (high title pool). Wilcoxon test did not show significant differences for haemolysis (p=0.0186), lipaemia (p=0.0017) or bilirubin (p=0.004).
Conclusions: Total antibodies electrochemiluminescent test showed an excellent clinical sensitivity with RT-PCR performed 10 days before. Intra-and inter-imprecision were satisfactory and the assay resulted unaffected by the three classical endogenous interfering.
