Explore the vast range of titles available.

The haemagglutination inhibition assay (HAI) and the virus microneutralisation assay (MN) are long-established methods for quantifying antibodies against influenza viruses. Despite their widespread use, both assays require standardisation to improve inter-laboratory agreement in testing. The FLUCOP consortium aims to develop a toolbox of standardised serology assays for seasonal influenza. Building upon previous collaborative studies to harmonise the HAI, in this study the FLUCOP consortium carried out a head-to-head comparison of harmonised HAI and MN protocols to better understand the relationship between HAI and MN titres, and the impact of assay harmonisation and standardisation on inter-laboratory variability and agreement between these methods. In this paper, we present two large international collaborative studies testing harmonised HAI and MN protocols across 10 participating laboratories. In the first, we expanded on previously published work, carrying out HAI testing using egg and cell isolated and propagated wild-type (WT) viruses in addition to high-growth reassortants typically used influenza vaccines strains using HAI. In the second we tested two MN protocols: an overnight ELISA-based format and a 3-5 day format, using reassortant viruses and a WT H3N2 cell isolated virus. As serum panels tested in both studies included many overlapping samples, we were able to look at the correlation of HAI and MN titres across different methods and for different influenza subtypes. We showed that the overnight ELISA and 3-5 day MN formats are not comparable, with titre ratios varying across the dynamic range of the assay. However, the ELISA MN and HAI are comparable, and a conversion factor could possibly be calculated. In both studies, the impact of normalising using a study standard was investigated, and we showed that for almost every strain and assay format tested, normalisation significantly reduced inter-laboratory variation, supporting the continued development of antibody standards for seasonal influenza viruses. Normalisation had no impact on the correlation between overnight ELISA and 3-5 day MN formats.

The recent outbreak of a novel Coronavirus (SARS-CoV-2) and its rapid spread across the continents has generated an urgent need for assays to detect the neutralising activity of human sera or human monoclonal antibodies against SARS-CoV-2 spike protein and to evaluate the serological immunity in humans. Since the accessibility of live virus microneutralisation (MN) assays with SARS-CoV-2 is limited and requires enhanced bio-containment, the approach based on “pseudotyping” can be considered a useful complement to other serological assays. After fully characterising lentiviral pseudotypes bearing the SARS-CoV-2 spike protein, we employed them in pseudotype-based neutralisation assays in order to profile the neutralising activity of human serum samples from an Italian sero-epidemiological study. The results obtained with pseudotype-based neutralisation assays mirrored those obtained when the same panel of sera was tested against the wild type virus, showing an evident convergence of the pseudotype-based neutralisation and MN results. The overall results lead to the conclusion that the pseudotype-based neutralisation assay is a valid alternative to using the wild-type strain, and although this system needs to be optimised and standardised, it can not only complement the classical serological methods, but also allows serological assessments to be made when other methods cannot be employed, especially in a human pandemic context.

•RSVpreF immunogenicity, safety, and tolerability were evaluated in healthy adults.•Equivalence in immunogenicity was assessed across 3 RSVpreF lots.•All 3 RSVpreF lots elicited strong immune responses 1 month after vaccination.•Lot-pair comparisons met the 1.5-fold equivalence criterion for both RSV A and RSV B.•RSVpreF single doses were safe and well tolerated, with safety similar across lots. Bivalent RSV prefusion F subunit vaccine (RSVpreF), comprised of equal quantities of stabilized prefusion F antigens from the major circulating subgroups (RSV A, RSV B), is licensed for prevention of RSV-associated lower respiratory tract illness (LRTI) in older adults and for maternal vaccination for prevention of RSV-associated LRTI in infants. To support licensure and large-scale manufacturing, this lot consistency study was conducted to demonstrate equivalence in immunogenicity across 3 RSVpreF lots. This phase 3, multicenter, parallel-group, placebo-controlled, randomized (1:1:1:1), double-blind study evaluated immunogenicity, safety, and tolerability of RSVpreF in healthy 18–49-year-old adults. Participants received a single 120-µg injection of 1 of 3RSVpreF lots or placebo. Geometric mean ratio (GMR) of RSV serum 50 % neutralizing geometric mean titers obtained 1 month after vaccination were compared between each vaccine lot for RSV A and RSV B, separately. Equivalence between lots was defined using a 1.5-fold criterion (GMR 95 % CIs for every lot pair within the 0.667–1.5 interval). Safety and tolerability were assessed. Of 992participants vaccinated, 948 were included in the evaluable immunogenicity population. All 3 RSVpreF lots elicited strong immune responses, meeting the 1.5-fold equivalence criterion for all between-lot comparisons for both RSV A and RSV B. Across the 3 lots, RSV A and RSV B 50 % neutralizing geometric mean titers substantially increased from baseline (RSV A, 1671–1795; RSV B 1358–1429) to 1 month after RSVpreF vaccination (RSV A, 24,131–25,238; RSV B, 19,238–21,702), corresponding to ≥14-fold increases in 50 % neutralizing titers for both RSV A and RSV B from before to 1 month after vaccination. Single doses of RSVpreF were safe and well tolerated, with similar safety profiles across the 3 RSVpreF lots. These findings support the reproducibility of RSVpreF vaccine manufacturing with similar safety and reactogenicity profiles (NCT05096208).

Background Influenza remains a significant threat to human and animal health. Assessing serological protection against influenza has relied upon haemagglutinin inhibition (HAI) assays, which are used to gauge existing immune landscapes, seasonal vaccine decisions and in systems vaccinology studies. HAI assays were first described in the 1940s. Here, we adapt our high‐throughput live virus microneutralisation (LV‐N) assay for SARS‐CoV‐2, benchmark against HAI assays, and report serological vaccine responsiveness in a cohort of older (> 65 yo) community dwelling adults. Methods Influenza‐specific antibody responses were assessed in 73 individuals, before and after receipt of the adjuvanted 2021–22 Northern Hemisphere quadrivalent vaccine. We performed both HAI and LV‐N assays against all four viruses represented in the vaccine [A/Cambodia/e0826360/2020 (H3N2), IVR‐215 (A/Victoria/2570/2019‐like) (H1N1)pdm09, B/Phuket/3073/2013 (B/Yamagata lineage), B/Washington/02/2019 (B/Victoria lineage)], using sera drawn before vaccination [range: d‐82 to d‐5], and days 7 [d6–10] and 181 [d156–200] after vaccination. We compared serological responses within each assay and between assays. Results Both the traditional HAI assay and our high‐throughput live virus microneutralisation identified vaccine‐induced boosts in antibody titres. We found population‐level concordance between the two assays (Spearman's correlation coefficient range 0.49–0.88; all p ≤ 1.4 × 10−5). The improved granularity of microneutralisation was better able to estimate fold changes of responses and quantify the inhibitory effect of pre‐existing antibody. Conclusions Our high‐throughput method offers an alternative approach to assess influenza‐specific serological responses with improved resolution, with the potential to improve the annual assessment of existing antibody landscapes, to improve new vaccine strain evaluation, and to offer a step‐change in systems vaccinology, and a facet of laboratory‐based pandemic preparedness.