The authors would like to thank Patrick Murray and Alex van Belkum for their generous and always positive support

The authors would like to thank Patrick Murray and Alex van Belkum for their generous and always positive support. study was performed in a retrospective multi-centric evaluation on aliquots of 328 nasopharyngeal samples. COVID-19 Ag Respi-Strip results were compared with Cadherin Peptide, avian qRT-PCR as golden standard for COVID-19 diagnostics. Results: In the analytical overall performance study, the reproducibility showed a between-observer disagreement of 1 1.7%, a robustness of 98%, an overall satisfying user friendliness and no cross-reactivity with other virus-infected nasopharyngeal samples. In the clinical performance study performed in three different clinical laboratories during the ascendant phase of the epidemiological curve, we found an overall sensitivity and specificity of 57.6 and 99.5%, respectively with an accuracy of 82.6%. The cut-off of the ICT was found at CT 22. User-friendliness analysis and risk management assessment through Ishikawa diagram demonstrate that COVID-19 Ag Respi-Strip may be implemented in clinical laboratories according to biosafety recommendations. Conclusion: The COVID-19 Ag Respi-Strip represents a promising quick SARS-CoV-2 antigen assay for the first-line diagnosis of COVID-19 in 15 min at the peak of the pandemic. Its role in the proposed diagnostic algorithm is usually complementary to the currently-used molecular techniques. diagnostic (IVD) medical devices (10), the Scandinavian SKUP-protocol (11) utilized for the validation of qualitative assessments and the clinical performance obtained with a multi-centric retrospective study. In addition, we reflect on the risk management and the conditions to be fulfilled before implementation as a point-of-care test (POCT) outside the hospital. Materials and Methods Development of the COVID-19 Ag Respi-Strip Antibodies and Antigen Eleven antibodies (designed A to K) (12) were coated at numerous concentrations on nitrocellulose (Advanced Microdevices, India) with antibodies A to J coupled to colloidal platinum beads (NanoQ, Belgium). Recombinant SARS-CoV nucleoprotein (recNP) preparation was obtained as explained previously (12) and was coated on a nitrocellulose membrane or conjugated on colloidal platinum nanoparticles. Recombinant his-tagged SARS-CoV-2 NP (recNP-2) has been produced in insect cells and purified (2-step purification), with a final purity 90% (Genscript, Leiden, NL). COVID-19 Ag Respi-Strip The ICT strip Rabbit Polyclonal to Tau consists of nitrocellulose laminated on a plastic backing, with colloidal-gold conjugates being dried on a conjugate pad (Ahlstrom-Munksj?, France) overlapping the bottom of the nitrocellulose. For preliminary direct detection, SARS-CoV and SARS-CoV-2 NPs were coated at 100 g/mL, and gold-labeled antibodies were deposited at 0.85 l/mm at 3 OD530 nm. The mean diameter of the gold nanoparticles is usually 40 nm. For the COVID-19 Ag Respi-Strip test, monoclonal antibodies directed against the SARS-CoV and SARS-CoV-2 highly conserved nucleoprotein antigen are coated around the nitrocellulose. Another monoclonal antibody is usually conjugated Cadherin Peptide, avian to colloidal platinum nanoparticles (mAb-gold nanoparticle). The conjugate is usually immobilized around the conjugate pad. The volume of the mAb-gold nanoparticle used in the conjugate release pad is usually 3.36 l per test (0.84 l per mm, while the strips are 4 mm wide). During the development, assessments analyzing the antibody reactivity and intensity were performed using serial dilutions of SARS-CoV-2 in a final volume of 300 l of buffer (data Cadherin Peptide, avian not shown here; cfr. Supplementary Furniture 2, 3). The results were decided after 15 min. For the experiments during development, the buffer volume was 300 l, while the buffer volume for the final test has been set at 200 l. During analytical and clinical overall performance studies, the final test used 200 l of buffer volume. The standard operating procedure for the COVID-19 Ag Respi-Strip is as follows (Physique 1): Transfer 100 L of a nasopharyngeal sample [nasopharyngeal aspirates (NPA), nasopharyngeal washes or nasal/nasopharyngeal swabs (NPS)] in the collection tube. Add 100 L of the LY-S dilution buffer to reach a dilution ratio of 1 1:2. The LY-S dilution buffer consists of TRIS buffer made up of EDTA, NaN3 ( 0.1%), Cadherin Peptide, avian a detergent and blocking brokers. Cap the tube with the stopper. Stir thoroughly to homogenize Cadherin Peptide, avian the solution. Open the tube. Immerse the strip in the.