reviewed the final draft of this article and provided logistic support

reviewed the final draft of this article and provided logistic support. is intended to be a core element in the global fight against the COVID-19 pandemic. Therefore, a variety of vaccines were approved rapidly. The protective role of mRNA-based SARS-CoV-2 BNT162b2 vaccine, as introduced by BioNTech/Pfizer, has been endorsed by several studies [1,2]. The rapidly increasing number of COVID-19 cases among the previously vaccinated individuals has led to the addition of a TSPAN3 third dose of vaccines as a booster [3,4].The immediate immune response after the third dose has been evaluated by a wide range of clinical trials [5,6,7,8]. Until now, there has been a lack of best clinical evidence about the durability and long-term impact of the immune response that is invariably induced by the third dose. Foregoing in view, we assessed the anti-spike-IgG-antibody-titers and neutralizing antibodies, 4 and 11 weeks after the third dose of BNT162b2 as well as 11 weeks after the administration of the second dose of BNT162b2 within a previously elaborated study cohort. == 2. Materials and Methods == == 2.1. Study Population == We included employees of the Hospital Reinbek St. Adolf-Stift, a German secondary care hospital to the longitudinal ProCoV-study, established in April 2020 [9]. In the follow-up analysis, all participants with a homogeneous prime-boost-protocol, who had received three IACS-8968 S-enantiomer doses of BNT162b2, were invited to participate in this study. Participants with a previous self-reported SARS-CoV-2-contamination within the last six months before the last blood drawn were excluded. All participants provided a blood sample 11 weeks after the second dose of vaccinate and then 4 weeks after the third dose between November and December 2021. Subsequently, in January 2022, a follow-up blood sample was drawn 11 weeks after the third dose. The study was approved in April 2020 by the ethics committee of the medical association Schleswig-Holstein, Germany and all participants provided written and informed consent prior to inclusion. This trial was prospectively registered in the German Clinical Trial Register (DRKS00021270). == 2.2. Anti-SARS-CoV-2-IgG Antibodies == Anti-SARS-CoV-2-IgG antibodies were measured using the anti-SARS-CoV-2- assay (IgG) from Abbott (Chicago, IL, USA) and the values were quantitatively expressed in binding-antibody-units per mL (BAU/mL). Values below 7.1 BAU/mL were determined seronegative whereas values above 7.1 BAU/mL were graded positive, as mentioned by the manufacturer. == 2.3. Surrogate Virus Neutralization Test == To detect neutralizing antibodies, a novel enzyme-linked immunosorbent assay (ELISA)-based IACS-8968 S-enantiomer surrogate virus neutralization test was used instead IACS-8968 S-enantiomer of a cell-culture based neutralization test, as this showed a good correlation and can easily be performed in routine laboratories [10]. With this, neutralizing anti-SARS-CoV-2 antibodies were detected according to the instructions of the manufacturer (NeutraLISA SARS-CoV-2 Neutralization Antibody Detection KIT (Euroimmun, Lbeck, Germany)). In brief, controls and plasma samples were diluted 1:10 and subsequently mixed with HRP-conjugated RBD. After incubation (37 C, 30 min) samples were transferred to a 96-well ELISA-plate precoated with recombinant ACE2 antigens. After incubating the plate (15 min, 37 C) and removing the supernatant, a substrate solution was added to each well. The reaction was stopped by adding a stop-solution to each well. Finally, adsorption was read at 450 nm. Results are presented in percentages of binding inhibition using the following formula: Binding inhibition [%] = (1(OD450 (sample)/OD450 (blank)) 100. Values above 35% were considered positive, between 20% and 35% equivocal and those below 20% were graded negative in accordance to the manufacturers instructions. == 2.4. Statistical Analysis == The statistical analysis was done using IACS-8968 S-enantiomer the IBM SPSS Statistics Version 25 (IBM Co., Armonk, NY, USA) and IACS-8968 S-enantiomer GraphPad Prism 9 (GraphPad Software, CA, USA). All variables were presented as means with standard deviation and medians with interquartile range. Categorical variables were shown as numbers with.