WizDx™ COVID-19 CrystalMix™ Multi PCR Kit INTENDED USE WizDx™ COVID-19 CrystalMix™ Multi PCR Kit is intended to be used to achieve qualitative detection of Severe acute respiratory syndrome coronavirus, (SARS-CoV-2) viral RNA extracted from nasopharyngeal swabs, oropharyngeal swabs, and sputum from patients. WizDx™ COVID-19 CrystalMix™ Multi PCR Kit combines all the reagents necessary for successful one-step real-time RT-PCR in a convenient individually aliquot and lyophilized in 8-strip qPCR tube. WizDx™ COVID-19 CrystalMix™ Multi PCR Kit uses dual-labeled hydrolysis probes that target two distinct regions in RdRP (RNA-dependent RNA polymerase) gene and E (Envelop) gene of SARS-CoV-2 genome. The detection probe for RdRP gene is labeled with FAM and the probe for E gene is labeled with HEX reporter dye. • Store at 4 ~ 25°C, Do not freeze. • Expires 12 months from date of manufacture. • Do not use once the cone-shape mix shrinks as dot-form. KIT CONTENTS Component COVID-19 CrystalMix COVID-19 Positive control * RNase-Free water * Amount 96 tubes 1 vial 1 vial * Before using the positive control, add 200μl of RNase-Free water and dissolve sufficiently before use. * Positive control & RNase-Free water are shipped at room temperature. After receipt, please store in refrigerated or frozen. REAGENT AND EQUIPMENT TO BE SUPPLIED BY THE USER • • • • CFX-96 Real-Time PCR System (Bio-Rad Laboratories, Inc.) Micropipette & sterile pipette tips Vortex mixer & microcentrifuge Protective ware and Disposable gloves TEST SAMPLE • Sputum, oropharyngeal swab, and nasopharyngeal swab samples SAMPLE COLLECTION, STORAGE AND TRANSPORTATION • Specimen collection assays are not provided with the assay. Refer to CDC guidelines for specimen collection and storage at: https://www.cdc.gov/coronavirus/2019-nCoV/guidelines-clinical-spec imens.html QUALITY CONTROL In accordance with Wizbiosolutions Inc. ISO 13485-certified Quality Management System, each lot WizDx™ COVID-19 CrystalMix™ Multi PCR Kit is tested against predetermined specifications to ensure consistent product quality. Carefully read this instruction before starting the procedure. Only use by authorized healthcare professionals. Use only for in vitro diagnostic purpose. Do not use any reagent after the expiration date. Always wear personal protective equipment (gloves and a mask) when handling biohazardous agents in compliance with national regulations. • Always use sterile, filtered pipette tips. • Take care of the handling of the specimen to minimize the risk of infection. • Dispose of waste in compliance with national or regional regulations after the test. PROTOCOL [Step 1] RNA Preparation The WizDx™ COVID-19 CrystalMix™ Multi PCR Kit has been validated with the Thermo Fisher RNA extraction kit (Catalog #: A42359). For the RNA extraction, please follow the manufacturer’s instructions. [Step 2] Prepare PCR Reaction 1) Prepare the PCR reaction as the following table. Component COVID-19 CrystalMix tube Sample (RNA) COVID-19 Positive Control RNase-Free Water Total * PTC : Positive template control, NTC : Non-template control ☞ To avoid contamination, close the cap immediately after placing the sample in the tube. 2) Vortex for 10 ~ 30sec. and briefly spin down for 5sec. 3) Insert the CrystalMix tube to the Real-Time PCR System. [Step 3] Run Real-time PCR 1) Prepare the Real-time PCR program set-up as the following table. Step cDNA Synthesis Initial Denaturation Denaturation Annealing (Probe detection) 2) Fluorescence probe and threshold setting as the following table. Fluoresence FAM HEX Cy5 Target RdRP E Internal Control [Step 4] Analysis of Results For interpretation, please refer to the Interpretation table. • Cut-off value of sample or positive control : Ct <40 Test COVID-19 detected Inconclusive COVID-19** No detected Invalid / Retest Note: * In case the target gene signal is strong, Cy5 (IPC) could be negative, and the result can be interpreted without IPC result. ** Depending on the sample condition, the sam

WizDx™ COVID-19 CrystalMix™ Multi PCR Kit EXAMPLE OF TEST RESULTS Fluorescence curve RdRP (FAM) PERFORMANCE EVALUATION The results for the COVID-19 assay performance evaluation have been generated on the CFX96™ Real-Time PCR detection system (Bio-Rad Laboratories, Inc.) with additional testing for analytical sensitivity. 1. Limit of Detection (LoD) To analyze the “Limit of Detection (LoD)” of WizDx™ COVID 19 CrystalMix™ Multi PCR Kit, a study was performed using SARS-CoV-2 RNA reference material, provided by Korea Research Institute of Standards and Science (KRISS) with serial dilution. Prepared...

WizDx™ COVID-19 CrystalMix™ Multi PCR Kit 3. Cross-reactivity To prevent the cross-reaction of the WizDx™ COVID-19 CrystalMix™ Multi PCR Kit, specific primers and probes to the RdRp and E were designed and produced. And cross-reactivity was assessed using the sequence information of viruses associated with respiratory disease registered in GenBank (http://www.ncbi.nlm.nih.gov/). Using the WizDx™ COVID-19 CrystalMix™ Multi PCR Kit, each of the respiratory virus species, resident flora, and other strains were tested. The list of viruses and strains used for the cross-reaction is shown in the table...

WizDx™ COVID-19 CrystalMix™ Multi PCR Kit TROUBLESHOOTING GUIDE Observation Possible Reason No signal increase is observed, even with positive controls • Pipetting errors • Check for correct reaction setup. Repeat the PCR run. • No data acquisition programmed. Fluorescence intensity is too low • Incorrect detection channel has been chosen. • Check the cycle programs • Low initial amount of target nucleic acid • Increase the amount of sample nucleic acid. • Exchange all critical solutions. Negative control samples are positive • Carry-over contamination. • Repeat the complete experiment with fresh...