Oropharyngeal and nasopharyngeal swabs were obtained from all participants for baseline PCR testing with a Bio-Speedy Direct RT-qPCR SARS-CoV-2 detection kit (Bioeksen, Istanbul, Turkey) on a Bio-Rad CFX96 Touch platform (Hercules, CA, USA), and serum total SARS-CoV-2 antibody testing was done. The ADVIA Centaur COV2T assay (Siemens Healthcare Diagnostics, Erlangen, Germany), a fully automated one-step antigen sandwich immunoassay using acridinium ester chemiluminescence technology, was used to detect total antibodies (IgG and IgM) against the SARS-CoV-2 spike protein receptor-binding domain (RBD) in serum samples. This assay is semiquantitative and has a lower detection threshold value (1 sample-to-cutoff ratio). All PCR and serum antibody tests were done at two central laboratories.

The study vaccine is an inactivated whole-virion vaccine with aluminium hydroxide as the adjuvant, prepared with a novel coronavirus (CZ02 strain) inoculated in African green monkey kidney cells (Vero cells). The inactivation process is done by adding β-propiolactone in the virus harvest fluid at a ratio of 1:4000 and inactivating at 2–8°C for 12–24 h. One dose of COVID-19 vaccine contains 3 μg of SARS-CoV-2 virion in a 0·5 mL aqueous suspension for injection with 0·45 mg/mL of aluminium. The placebo contained all ingredients except the inactivated virus, in prefilled syringes. The injections were given in two doses, 14 days apart, intramuscularly in the deltoid muscle. As the placebo and study vaccine looked exactly the same, they were administered by staff masked to group allocation. Details of the procedures on visit dates and the pharmacological properties of the investigational product are provided in the appendix (pp 1–2).

Symptom-based active surveillance was done to detect participants with symptoms suggestive of COVID-19 during follow-up (appendix pp 3–4). Anyone with at least one of the following symptoms for 2 days or more underwent PCR testing: fever or chills; cough; dyspnoea; fatigue; muscle or body pain; headache; new loss of sense of smell or change in taste; sore throat; nasal congestion or rhinorrhoea; nausea or vomiting; and diarrhoea. Cases of SARS-CoV-2 infection were classified according to the scale of clinical progression proposed by WHO.11 Clinical outcomes were assessed in a blinded manner.

Sampling for immunogenicity analyses was planned in a subgroup of volunteers selected sequentially. As the immunogenicity and T-cell response analyses are ongoing, we only report the initial results of the anti-RBD antibody tests and neutralising antibody assays gathered at least 14 days after the second dose of vaccine or placebo. Virus neutralisation assays were done in an in-house microtitre plate, as described by Hanifehnezhad and colleagues.12 Five-fold diluted serum samples, starting from 1:5, were mixed with an equal volume of 100 median tissue culture infectious dose of SARS-CoV-2 Ank1 isolate (1:10 000) in quadruplicate and incubated for 1 h at 37°C for neutralisation. The serum–virus mixtures were subsequently inoculated onto 90% confluent Vero E6 cells grown in 96-well plates. The assay was evaluated via inverted microscope when a 100% cytopathic effect was observed in the virus control wells. Reciprocals of serum dilutions inhibiting at least 50% of virus infectivity were expressed as mean antibody titre (SN50).

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