DOI: 10.5061/dryad.ghx3ffbsw
title: Data for: Evaluation of antibody kinetics and durability in health individuals vaccinated with inactivated COVID-19 vaccine (CoronaVac): a cross-sectional and cohort study in Zhejiang, China

description: Background: Although inactivated COVID-19 vaccines are proven to be safe
 and effective in the general population, the dynamic response and duration
 of antibodies after vaccination in the real world should be further
 assessed. Methods: We enrolled 1067 volunteers who had been vaccinated
 with one or two doses of CoronaVac in Zhejiang Province, China. Another 90
 healthy adults without previous vaccinations were recruited and vaccinated
 with three doses of CoronaVac, 28 days and 6 months apart. Serum samples
 were collected from multiple timepoints and analyzed for specific IgM/IgG
 and neutralizing antibodies (NAbs) for immunogenicity evaluation. Antibody
 responses to the Delta and Omicron variants were measured by
 pseudovirus-based neutralization tests. Results: Our results revealed that
 binding antibody IgM peaked 14–28 days after one dose of CoronaVac, while
 IgG and NAbs peaked approximately 1 month after the second dose and then
 declined slightly over time. Antibody responses had waned by month 6 after
 vaccination and became undetectable in the majority of individuals at 12
 months. Levels of NAbs to live SARS-CoV-2 were correlated with
 anti-SARS-CoV-2 IgG and NAbs to pseudovirus, but not IgM. Homologous
 booster around 6 months after primary vaccination activated anamnestic
 immunity and raised NAbs 25.5-fold. The neutralized fraction subsequently
 rose to 36.0% for Delta (p=0.03) and 4.3% for Omicron (p=0.004), and the
 response rate for Omicron rose from 7.9% (7/89) to 17.8% (16/90).
 Conclusions: Two doses of CoronaVac vaccine resulted in limited protection
 over a short duration. The inactivated vaccine booster can reverse the
 decrease of antibody levels to prime strain, but it does not elicit potent
 neutralization against Omicron; therefore, the optimization of booster
 procedures is vital.
description: Study
 design and participants The cross-sectional investigation was conducted
 in five
 counties of
 Zhejiang
 Province, mainland China
 (Xihu, Yuecheng, Shangyu,
 Kaihua, and Longyou Districts), after nationwide COVID-19 vaccinations from May
 to October 2021. Potential participants aged 18–59 years who had no prior
 vaccinations or were vaccinated with one or two doses of CoronaVac
 (Sinovac Life Sciences, Beijing, China) were recruited from the community.
 Individuals with a history of infection with SARS-CoV-2 (based
 on epidemic surveillance
 system) or the use of
 blood products or immunosuppressive drugs were excluded. We randomly
 enrolled 1067 volunteers,
 including those on day 0
 (V-0, no vaccination), day 14 ± 2 (V1-14d), and day 28 ± 3 (V1-28d) after
 the first vaccine dose, and day 30 ± 3 (V2-1m), day 90 ± 7 (V2-3m), day
 180 ± 14 (V2-6m), day 270 ± 14 (V2-9m), and day 365 ± 30 (V2-12m) after
 the second dose and
 collected
 their venous blood samples (3–5 ml) to detect serum antibody
 levels (Figure 1A). This
 was not a longitudinal survey, as different subjects were enrolled at each
 point in time. We employed
 a questionnaire survey at blood drawing visits to gather demographic
 information. In the prospective cohort study, we recruited 90
 healthy adults aged 18–80 years from Jiaxing city, Zhejiang, in June 2021. The main exclusion criteria
 included previous or later SARS-CoV-2 infection; allergy to any ingredient
 included in the vaccine; those who had received any blood products or any
 research medicines or vaccines in the past month; those who had
 uncontrolled epilepsy or other serious neurological diseases, acute
 febrile disease, acute onset of chronic diseases, or uncontrolled severe
 chronic diseases; and those who were unable to comply with the study
 schedule. Subjects were administered 4 µg/0.5 mL of CoronaVac following a 3-shot vaccine schedule 28 days and
 6 months apart. Following that, venous blood (3–5 ml) was collected from
 recipients at five time points: day 0 (Pre-V, before vaccination), day 30 ± 3 (V2-1m), day 90 ± 7 (V2-3m), and day
 180 ± 14 (V2-6m) after the
 second dose, and day 30 ±
 3 (V3-1m) after the third
 dose (Figure
 1B). SARS-CoV-2-specific IgG and IgM
 assay The commercial detection kit iFlash-2019-nCoV
 NAb assay (Shenzhen YHLO Biotech Co. Ltd., Shenzhen, China) was employed
 to measure the levels of
 IgG and IgM against SARS-CoV-2 spike glycoprotein (S) and nucleocapsid
 protein (N) by chemiluminescence immunoassay. Briefly, serum samples were
 allowed to form a complex with SARS-CoV-2 S- and N-protein antigen-coated
 paramagnetic microparticles, then an acridinium-ester-labeled ACE2
 conjugate was added to competitively combine with the particles, forming
 another reaction mixture. The analyzer converted relative light units
 (RLUs) into an antibody titer (AU/mL) through a two-point calibration
 curve. An inverse relationship existed between the amount of SARS-CoV-2
 NAb in the sample and the RLUs detected by the iFlash optical system.
 According to the manufacturer, titers of ≥10.0 AU/mL and ≥1.0 AU/mL are
 considered positive (or reactive) for IgG and IgM, respectively. IgG and
 IgM against SARS-CoV-2 receptor binding domain (RBD) were detected using a
 commercial ELISA kit (Bioscience Biotech Co. Ltd., Chongqing, China). The positive cutoff values for RBD-specific IgG
 and IgM
 antibodies were defined as
 titers of ≥1.0
 AU/mL. All tests were
 performed according to the manufacturer’s protocols (Chan et al. 2021; Li et al.
 2021). Live virus
 neutralization antibody assays The levels of neutralizing antibodies to live
 SARS-CoV-2 were assessed by the reduction in the cytopathic effect (CPE)
 in Vero cells with infectious SARS-CoV-2 strain 19nCoVCDC-Tan-HB01 (HB01)
 in a BSL-3 laboratory(Zhang et al. 2021). Briefly, serum samples were heat-inactivated
 for 30 min at 56°C and successively diluted from 1:4 to the required
 concentration in a 2-fold series. An equal volume of challenge solution
 containing 100 TCID50 virus was added. After neutralization in a 37°C
 incubator for 2 h, a 1.5–2.5 × 10<sup>5</sup>/ml cell
 suspension was added to the wells. The CPE (cytopathic effect) on VeroE6
 cells was analyzed at 4-days post-infection. NT50 (50% neutralization
 titer, the reciprocal of the highest dilution protecting 50% of the cells
 from virus challenge) was used to show the neutralization titers. NT50
 above 1:4 was defined as positive. Pseudovirus-based neutralization
 test Serum samples were also quantified for their
 content of SARS-CoV-2-neutralizing antibodies to wildtype (Wuhan), Delta
 (B.1.617.2), and Omicron (B.1.1.529) using the pseudovirus-based virus
 neutralization test (Nie
 et al. 2020). Briefly,
 serum samples and a positive or negative reference sample were each
 diluted 50 times with phosphate-buffered saline combined with 50 µl of
 pseudovirus diluent per well in a 96-well plate. The mixed
 sample/pseudovirus was incubated at 37°C and 5% CO<sub>2</sub>
 for 1 h. A 2 × 10<sup>5</sup>/ml BHK-21-ACE2 cell suspension
 was added to each well of the plate containing the sample/pseudovirus
 mixture, then the plate was incubated in a 37°C and 5%
 CO<sub>2</sub> cell incubator for 48 h. Finally, the number of
 green-fluorescence-protein-positive cells per well was read with a porous
 plate imager (Tecan, Shanghai, SparkCyto). The results were determined by
 comparing the neutralized
 fraction using the
 following calculation: (1 – (fluorescence value of each well/average virus
 control value)) × 100%(Karaba et al. 2022). At least 4 wells were left blank for
 calibration to 0% inhibition. The conversion of neutralized fraction to serum titer was shown in
 Table
 S2. Statistical analysis
 Sex, age, BMI, and other clinical
 characteristics were collected for each vaccination recipient. We used the
 medians and interquartile ranges (IQR) for age, and numbers (percentages)
 for categorical variables. Specific binding antibodies against SARS-CoV-2
 (IgG, IgM) and neutralized fraction of SARS-CoV-2-neutralizing antibodies
 are presented as mean ± standard error (SEM). Neutralizing antibodies are
 presented as geometric mean titers (GMT), and their 95% confidence
 interval (CI) was calculated with Student’s t distribution on
 log-transformed data and then back-transformed. Comparisons of titer-level
 differences between the two groups were performed using the paired
 Student's t-test. One-way analysis of variance (one-way ANOVA) was
 used to analyze the differences between the mean values at different
 timepoints. Correlations between NAb titers, neutralized fraction, and
 IgG/IgM levels were evaluated by Pearson’s correlation coefficient.
 Statistical tests were two-sided, and we considered p-values of less than
 0.05 as statistically significant. All statistical analyses were conducted
 in SPSS 18.0 (IBM Corporation, Armonk, NY, USA) and GraphPad Prism 9 (San
 Diego CA, USA).