Analytical performance evaluation of the EDELVITAL neutralizing antibody test in COVID-19-vaccinated staff of the University Dental Clinic Vienna

The assessment of immune responses plays a central role in evaluating the degree of protection after SARS-CoV-2 vaccination. The Abbott Alinity-I SARS-CoV-2 IgG II Quant Assay is widely used as a laboratory-based reference method for the quantitative assessment of anti-SARS-CoV-2 antibodies. In 2021, numerous rapid point-of-care antibody assays entered the market; however, their analytical performance and comparability with established automated platforms have not been comprehensively evaluated. The EDELVITAL fluorescence immunochromatographic assay is a fluorescence-based rapid test designed to assess neutralizing antibodies at the point of care. Early data demonstrated substantially increased infection risks among frontline healthcare personnel compared with the general population. In this context, reliable information on post-vaccination humoral immunity was of particular occupational and public health relevance. Given the limited independent validation data available for rapid assays, at this time, it was uncertain whether they reliably detected an adequate post-vaccination antibody response in comparison to gold standard methods. Therefore, this study aimed to evaluate the performance of a rapid point-of-care neutralizing antibody assay in vaccinated healthcare workers and to assess its concordance with the established Abbott Alinity I IgG assay. Because the two assays measure different aspects of the humoral immune response, the study also explored the potential complementary value of rapid neutralizing antibody testing in occupational health settings. A total of 334 serum samples from 291 COVID-19-vaccinated individuals were analyzed. IgG antibodies were quantified using the automated Abbott Alinity I CMIA, while neutralizing antibodies were measured using the EDELVITAL fluorescence immunochromatography test, calibrated to the WHO International Standard NIBSC 20/136 and SOP. Correlation and diagnostic analyses were performed. The two assays showed a high level of agreement (95.5% of the EDELVITAL neutralizing assay results agreed with those from the Abbott Alinity I system) when it came to detecting positive results. The rapid point-of-care device presented reproducible neutralizing antibody results (positive/negative) within 15–20 min, aligning closely with the laboratory-based Abbott measurements. The EDELVITAL fluorescence immunochromatographic assay showed analytical comparability with the Abbott Alinity I SARS-CoV-2 IgG II Quant Assay and may represent a practical complementary tool for antibody monitoring in healthcare workers. The rapid availability of its results can facilitate timely decisions regarding booster vaccination and occupational health management.

Following its first detection in Wuhan, China, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), associated with the acute respiratory syndrome coronavirus disease 2019 (COVID-19), has challenged healthcare systems worldwide. Coronavirus disease 2019 (COVID-19), was first identified in Wuhan, China, in late 2019. Following its rapid global spread, the World Health Organization (WHO) declared COVID-19 a Public Health Emergency of International Concern on 30 January 2020 and subsequently characterized the outbreak as a pandemic on 11 March 2020. The pandemic resulted in an unprecedented global public health challenge and prompted extensive efforts to develop diagnostic tools, vaccines, and strategies for monitoring immune responses (1, 2). Beyond its direct health consequences, the COVID-19 pandemic profoundly affected daily life, social interactions, and working environments worldwide. While many professions transitioned to remote work and home-office arrangements (3), healthcare professionals remained at the forefront of patient care and were therefore at increased risk of occupational exposure to SARS-CoV-2. This risk was particularly pronounced in dental settings, where aerosol-generating procedures and close patient contact substantially increased the potential for viral transmission. Early epidemiological data demonstrated substantially increased infection risks among frontline healthcare personnel compared with the general population (4), highlighting the particular vulnerability of specific occupational groups during the pandemic. In response to the rapid global spread of the virus, vaccines were swiftly developed and deployed (5), although their accelerated introduction was accompanied by concerns and mistrust among certain segments of the population (6, 7). An individualized immunization strategy could help to conserve already-limited financial and human resources, hospital capacity, and essential healthcare infrastructure. Considering that there were shortages of hospital beds and medical personnel during the pandemic, even in highly developed healthcare systems (8), such an approach could contribute to better resource allocation and preparedness (9).

Antibody-based immunity—in particular, the formation of spike protein-specific immunoglobulin G (IgG) and neutralizing antibodies (nAbs)—plays a central role in protecting against symptomatic infections and severe disease progression (9–11). Long-term studies have shown that a stable immune response persists for several months after natural infection (10–13) and/or vaccination, with the neutralizing antibody titers highly predictive of the degree of clinical protection (10, 11). Antibody detection has therefore become a crucial tool in vaccine research and immune monitoring. In addition to modeling studies linking neutralizing antibody titers to clinical protection, comparative analyses across multiple COVID-19 vaccine trials have demonstrated a strong correlation between both binding (Spike-specific IgG) and neutralizing antibody levels and observed vaccine efficacy, supporting antibodies as robust correlates of protection against SARS-CoV-2 (14). After vaccination with mRNA-based vaccines (BNT162b2, mRNA-1273), individuals show robust IgG and nAb responses after second vaccination, but their strength and duration vary depending on age and time (14–17). Therefore, it is urgent to identify a reliable correlate of protection for SARS-CoV-2 vaccines to guide vaccination strategies and immune monitoring (18), particularly in highly exposed populations such as healthcare workers. In this context, serological testing is also gaining importance in everyday clinical practice, as it provides information on individual immune status and therefore the potential need for boosters.

Various serological methods are available for the quantitative assessment of the immune response to vaccination. The Abbott Alinity I SARS-CoV-2 IgG II Quant Assay is a fully automated chemiluminescent micro particle immunoassay (CMIA) that determines spike RBD-specific IgG in BAU/mL (17) and is calibrated to the WHO standard NIBSC 20/136 (19). This system is used in many routine clinical laboratories and offers high analytical precision with high sample throughput. In contrast, the EDELVITAL fluorescence immunochromatography test is based on a competitive rapid test principle that measures blocking of the ACE2-RBD interaction and thus indirectly quantifies the functional neutralization capacity (20). Since binding antibody assays and neutralization assays assess distinct aspects of the humoral immune response, they should be considered complementary rather than competing approaches for immune monitoring. The test is performed using a portable fluorescence reader (AF-100S), and its results are given in internationally standardized IU/mL units (WHO reference 20/136) (19) within 15–20 min. A key advantage of this system is its suitability for clinical use (Figure 1), as it enables straightforward, near-patient quantitative assessment of neutralizing antibody activity without the need for a specialized laboratory. This is particularly practical for occupational health applications, including among healthcare workers, as well as for booster vaccination planning and immunosuppressed patients. It can also be used with a finger prick test.

Previous comparative studies have predominantly evaluated the analytical performance of individual serological assays under laboratory or study-specific conditions. Serological assays played an important role in monitoring SARS-CoV-2 exposure and vaccine-induced immune responses throughout the pandemic. Among these, enzyme-linked immunosorbent assays (ELISAs) became widely used because of their accessibility, standardized laboratory implementation, and ability to quantify specific antibody responses (21). However, ELISA-based methods generally require laboratory infrastructure and longer processing times, which may limit their applicability in point-of-care or occupational health settings. Levin et al. demonstrated in a large prospective cohort that anti-Spike IgG and nAb titers decline progressively over six months following two-dose mRNA vaccination, with neutralizing antibodies showing a more pronounced early decrease and significantly lower levels in older and immunosuppressed individuals (12). Nouanesengsy et al. reported that hybrid immunity (prior infection followed by vaccination) induces robust and broader neutralizing responses, characterized by a strong antibody boost after the first vaccine dose and enhanced variant neutralization compared with infection alone, while additional doses conferred only limited incremental increases (13). While both studies reported comparable patterns in IgG and nAb decreases, Takheaw et al. reported that, while anti-RBD IgG levels correlated well with neutralizing activity against the ancestral strain and earlier variants, this relationship appeared less consistent for the Omicron variant, suggesting that the predictive value of binding antibody levels may vary depending on the circulating viral strain (22). Investigations of automated quantitative assays primarily focused on assay precision, sensitivity, and inter-method comparability, without addressing their use within routine clinical workflows (23, 24). Conversely, studies assessing rapid or lateral flow-based tests largely examined their feasibility or analytical characteristics as stand-alone tools, often outside real-world clinical settings (24, 25). Importantly, the potential complementary use of automated laboratory assays and rapid near-patient testing within routine clinical practice has not been systematically evaluated. This lack of evidence regarding integrated testing approaches in everyday healthcare settings provided the rationale for the present study. In contrast to the abovementioned laboratory-based longitudinal investigations focused on two-dose vaccination (12) or hybrid immunity (13), our study specifically evaluates IgG and nAbs before and after a third mRNA vaccine dose in SARS-CoV-2-vaccinated staff of the University Dental Clinic Vienna. To verify a sufficient neutralizing antibody response against severe COVID-19 infection, a rapid point-of-care assay was used, thereby providing insight into booster-associated immunogenicity in a real-world setting. The present study aimed to investigate the relationship between quantitative anti-SARS-CoV-2 IgG measurements obtained with the Abbott Alinity I assay and neutralizing antibody measurements generated by the EDELVITAL fluorescence immunochromatographic assay. Furthermore, we explored the potential utility of rapid neutralizing antibody testing as a complementary tool for assessing vaccine-induced humoral immunity in healthcare workers and explored the clinical utility of rapid serological testing to support evidence-based “test-before-boost” decision strategies. Because age- and sex-specific distributions are reported to be crucial in immune responses (26, 27), these factors were also analyzed by comparing age group and male/female responses.

This prospective cohort was conducted at the University Dental Clinic of the Medical University of Vienna between July 2021 and March 2022. Data were obtained from routine clinical laboratory diagnostics performed as part of employee testing or after completion of the primary COVID-19 vaccination. During the study period, healthcare workers were subject to routine institutional SARS-CoV-2 screening measures established for occupational health and infection control purposes, including twice-weekly rapid antigen testing and confirmatory PCR testing when indicated. Individuals with documented SARS-CoV-2 infection were excluded from the study cohort. This approach was considered particularly relevant, as previous studies have demonstrated that SARS-CoV-2 IgG antibodies may arise from both vaccination and prior infection, potentially complicating the interpretation of serological findings in occupationally exposed healthcare populations (21, 28).

Sample collection and subject characteristics

After applying predefined inclusion (a minimum of two vaccine shots against SARS-CoV-2 within the last 4–8 months (BioNTech, Pfizer, Moderna), age 16+) and exclusion criteria (symptomatic for COVID-19 between sample collection, immunocompromised subjects, incomplete data, invalid samples), 334 serum samples of 291 individuals were eligible for analysis. Of these, 257 samples were obtained before (pre-booster) and 77 after (post-booster) the third vaccine dose. Paired samples from the same individual before and after the third vaccination were available in 43 cases. The demographic variables included sex, vaccination type, time since last dose, and predefined age groups (16–26, 27–35, 36–50, and >50 years).

This study focuses on a reliable binary (yes/no) assessment of a sufficient (prevent severe COVID-19 infection) vaccine-induced immune response in a clinical setting through monitoring binding IgG and nAbs. Blood samples were taken 4–8 months after primary immunization (two shots) and again three weeks after the third booster vaccination, and IgG and nAb levels were analyzed. The correlations of IgG and nAb responses with infections and vaccines are reported (10–16), allowing for a correlation analysis in our results between the increase in IgG and nAb antibodies. The antibody responses were also related to differences by sex and age group.

The participants had received different primary immunization regimens based on the COVID-19 vaccines authorized at the time. The mRNA-based vaccines Comirnaty (BNT162b2, BioNTech/Pfizer) and mRNA-1273 (Moderna) were administered in two-dose schedules; both formulations contain nucleoside-modified mRNA encoding the SARS-CoV-2 spike protein, encapsulated in lipid nanoparticles to ensure cellular uptake and protection of the RNA (9). Blood samples were collected intravenously using 9 mL VACUETTE ® CAT Serum Sep. Clot Activator tubes 4–8 months after the primary immunization (before the third booster), and three weeks after the third booster.

Analytical methods

IgG concentrations were quantified with the Abbott Alinity I SARS-CoV-2 IgG assay in the Austrian Agency for Health and Food Safety (AGES) laboratories. Neutralizing antibody levels were determined using a fluorescence immunochromatographic assay (EDELVITAL rapid test system) at the Core Facility for Oral Microbiology and Hygiene, Dental Clinic, Medical University of Vienna.

Abbott Alinity I SARS-CoV-2 IgG II quant assay

The IgG antibodies were quantified using the Abbott Alinity I SARS-CoV-2 IgG II Quant Assay (Abbott Diagnostics, Chicago, IL, USA). This test is a chemiluminescent microparticle immunoassay (CMIA) that features the recombinant receptor binding domain (RBD) antigen of the SARS-CoV-2 spike protein as the target antigen. Antibodies in the sample bind to RBD-coated microparticles. After adding an acridinium-labeled anti-human IgG conjugate, the chemiluminescence signal is measured in proportion to the antibody concentration (29).

Calibration: The assay is calibrated to the WHO standard NIBSC 20/136 (19). The results are reported in binding antibody units (BAU/mL).

Evaluation: Values ≥ 7.1 BAU/mL were considered positive (cut-off), according to the manufacturer’s instructions; measuring range: 2.98–5680 BAU/mL.

Analytical precision: Intra-assay variability ≤ 5%; inter-assay variability ≤ 6%.

Quality control: Daily internal control measurements with two calibrators and two quality controls according to the manufacturer’s recommendations.

This system enables high sample throughput.

Quality assurance

Commercial control materials provided by the manufacturer (Abbott SARS-CoV-2 IgG II Quant Control Kit) containing defined concentrations (2 positive, 1 negative) of anti-SARS-CoV-2 IgG antibodies were analyzed at least once every 24 hours on each day of assay use, with the first run of patient sample analysis. Patient results were only released when all the quality control results were within the manufacturer’s predefined acceptance limits. If the control results fell outside the predefined acceptance range, the patient results obtained in that run were considered invalid. The laboratory followed its internal quality control procedures, including troubleshooting and repeating the measurements. The assay was recalibrated if necessary. The software was updated multiple times (versions 3.2.1, 3.2.2, 3.2.2, and 3.2.3).

Additionally, the laboratory successfully participated in annual external quality assessment schemes (INSTAND e.V., Germany), demonstrating acceptable performance throughout the study period.

EDELVITAL fluorescence immunochromatography test

This test (AMEYA GmbH, Vienna, Austria) was used to determine neutralizing antibodies using competitive fluorescence immunochromatography, which measures the inhibition of binding between the viral RBD and the human ACE2 receptor.

Measuring principle: Serum samples are incubated with fluorescence-labeled RBD protein.

Neutralizing antibodies block the binding of ACE2 to the RBD, which reduces the intensity of the fluorescence signal in the test field.

The immunofluorescence reader (analyzer) provides the results in IU/mL.

According to the manufacturer’s (AMEYA) specifications, the initial samples were measured in arbitrary units per milliliter (U/mL). To comply with World Health Organization (WHO) standards and ensure comparability, all results were subsequently converted and reported in WHO binding antibody units per milliliter (IU/mL). The conversion was performed using the following equation:

U/mL = iU/mL × 0.142.

This conversion factor was established by the manufacturer based on internal validation studies conducted in accordance with the WHO International Standard for anti–SARS-CoV-2 immunoglobulin (NIBSC Code 20/136) (19).

Quality assurance

The Fluorescence Immunochromatography Test was calibrated before each run with a positive and negative control. These controls were provided with the Test system. Patient results were only released when all the quality control results were within the manufacturer’s predefined acceptance limits. If the control results fell outside the predefined acceptance range, the patient results obtained in that run were considered invalid.

Data processing and statistical analysis

The raw data were screened to remove duplicates and non-eligible records. Data cleaning and preparation were performed in Microsoft Excel 365, while statistical analyses were conducted using IBM SPSS Statistics 28. Figures and tables were generated using both software packages. The final dataset was stratified by sex and predefined age group (16–26 years, 27–35 years, 36–50 years, >50 years). Descriptive statistics (mean, median, and quartiles) were calculated for the total cohort and all subgroups.

Group comparisons

Standard parametric procedures were applied as follows:

Sex and age group differences

Independent-sample t-tests were used to compare the mean antibody levels between males and females. Variance homogeneity was evaluated with Levine’s test; in cases of unequal variances, the Welch t-test was applied.

One-way ANOVA was used to compare the mean antibody levels across the four age groups.

If the variances were homogeneous, Tukey’s post hoc test was applied.

If variance heterogeneity was present, Welch’s ANOVA with Games–Howell post hoc testing was used.

Booster effect and quantitative comparison

Changes in antibody levels before and after the third vaccine dose were assessed using paired-samples t-tests. To assess the correlation between the IgG and neutralizing antibody assays, Pearson’s coefficient of correlation and Spearman’s coefficient of correlation among the four assays were calculated.

Diagnostic performance (crosstab analysis)

A crosstab approach was applied to compare qualitative IgG and neutralizing antibody classifications. This enabled calculation of the agreement in detecting a sufficient antibody response between the EDELVITAL rapid test system relative to the Abbott Alinity-I platform.

To minimize the risk of false-negative results, samples yielding negative EDELVITAL measurements were re-analyzed up to three times before final classification. Abbott Alinity testing was performed in an accredited external laboratory under routine quality-controlled conditions and served as an established serological reference method. The concordance and discordance between both assays were subsequently evaluated by cross-tabulation analysis, allowing assessment of the agreement between EDELVITAL classifications and Abbott-based serological status.

All the statistical tests were two-sided with a significance level of p < 0.05. Normality and variance assumptions were systematically assessed to ensure the appropriate choice of parametric methods and to maintain methodological robustness (Supplementary Material).

Study population

A total of 334 serum samples from 291 vaccinated individuals were analyzed. Their average age was 35 ± 10 years (Figure 2), and 65.9% of the participants were female. Of the samples, 257 (77%) were taken after the second vaccine dose (pre-booster) and 77 (23%) after the third dose (post-booster); of these, 43 individuals had matching pre- and post-booster samples. All detailed analyses are available in the Supplementary Material (Block A: Descriptive statistics age and sex, Supplement Tab 1-3.

Sex- and age-dependent differences in booster response

Minor sex- and age-related differences in booster responses were observed. Men showed slightly lower neutralizing antibody levels than women prior to the third dose, while no other sex-dependent differences were detected.

Exploratory age-stratified analyses showed descriptive differences in antibody levels between age groups. Statistical testing indicated significant age-related differences only for EDELVITAL measurements before the third vaccine dose, whereas no statistically significant age-group differences were detected after booster vaccination. Therefore, age-group comparisons should be interpreted cautiously, particularly in view of the observed variability within groups. (Supplementarys Block A Tables 1, 2 and 3). Despite these limited effects IgG (Figure 3) and neutralizing antibody nAb (Figure 4) responses before and after the third (booster) vaccination are presented in separate graphical displays, with age groups included for descriptive stratification (Supplementary Block B: Comparison Sex and Age before and After 3rd vaccination: Figures 1–8 and Supplementary Tables 4-26.

Booster effect and quantitative correlation

In the analysis of the collective sample set, all participants tested positive for SARS-CoV-2 IgG antibodies both 4–8 months after completion of the primary vaccination series with two doses of the Pfizer/BioNTech vaccine and two weeks after receiving a third (booster) dose with the Moderna vaccine. The mean IgG concentration increased markedly after the booster vaccination, rising from 913.8 BAU/ml prior to the third dose to 4727.3 BAU/ml after the booster (Figure 3).

In contrast, the mean levels of neutralizing antibodies in the collective dataset were slightly lower after the booster dose compared to before the booster (70.2 IU/ml vs. 75.84 IU/ml) (Figure 4). Nevertheless, the qualitative classification of neutralizing antibodies remained largely unchanged. In total, 94.2% of the samples collected prior to the third vaccine dose and 100% of the samples collected after the booster dose tested positive for neutralizing antibodies.

The observed differences between IgG and neutralizing antibody responses should be interpreted in the context of the distinct biological parameters assessed by each assay. While the Abbott assay quantifies binding antibodies directed against the SARS-CoV-2 spike RBD, the EDELVITAL assay assesses functional neutralizing activity. Therefore, direct quantitative comparison between both measurements is inherently limited. Additional tables are provided in the Supplementary Material Block B, Figures 5-8, Tables 4-26.

A separate analysis of the paired-sample subset, consisting of samples obtained from the same individuals before and after booster vaccination, showed a different pattern. In the paired-sample subset, mean Abbott Alinity IgG concentrations increased from 114.4 BAU/ml before the third vaccine dose to 4716.7 BAU/ml after booster vaccination, corresponding to an approximately 41-fold increase in antibody levels (Figure 5).

In the same paired-sample subset, mean EDELVITAL measurements increased from 48.5 IU/ml before the third vaccine dose to 75.0 IU/ml after booster vaccination, corresponding to an approximately 56% increase in surrogate neutralizing antibody activity. Figure 6, Supplementary Material Block C: Difference before and after 3rd vaccination dose: Figures 9 and 10, Tables 27-29.

Correlation analyses were performed to examine the relationship between IgG antibody concentrations and neutralizing antibody (nAb) levels before and after the third vaccine dose. Pearson correlation analysis revealed a moderate positive correlation between IgG levels measured after the third dose and nAb levels measured prior to the booster (r = 0.574, p < 0.001) (Figure 7).

In addition, a significant positive correlation was observed between IgG and nAb levels prior to the third vaccine dose (r = 0.437, p < 0.001). A weaker but still statistically significant correlation was also detected between IgG levels measured before and after the booster vaccination (r = 0.320, p = 0.036).

These results indicate statistically significant associations between IgG concentrations and neutralizing antibody levels across the different sampling time points. The observed moderate correlations suggest that individuals with higher SARS-CoV-2 IgG concentrations generally also exhibit higher surrogate neutralizing antibody responses. However, the correlations were not sufficiently strong to indicate complete interchangeability of the assays, supporting the concept that Abbott Alinity and EDELVITAL capture related but distinct aspects of the vaccine-induced humoral immune response.

The Spearman correlation was additionally calculated to confirm the robustness of the associations (Figure 8).

Spearman’s rank correlation confirmed these findings, demonstrating a strong positive correlation between IgG and NAb levels before the third dose (rs = 0.631, p < 0.001). Additional significant associations were observed between IgG levels before and after the booster (rs = 0.382, p = 0.012) and between NAb levels prior to the third dose and IgG levels after the booster (rs = 0.528, p < 0.001). Overall, these results indicate consistent positive relationships between IgG and neutralizing antibody responses across time points. Supplementarys Block D: Correlations, Figures 11–16 and) Tables 30, 31.

Diagnostic performance (crosstab analysis)

To determine the efficacy of a labor-independent point-of-care device for detecting a sufficient antibody response to SARS-CoV-2 vaccination to prevent severe infection, a crosstab approach (Supplementary Material) was performed. Positive binding antibody detection was compared between the labor-based gold standard Abbott Alinity I SARS-CoV-2 IgG II Quant Assa y and rapid point-of-care system EDELVITAL Fluorescence Immunochromatography Test (Table 1).

Concordance between the IgG and nAb

The positive results of the EDELVITAL nAb assay were compared with the Abbott Alinity IgG assay results among all samples. The EDELVITAL nAb Test (EDELVITAL) detected 319 samples (95.5%) as positive (above the cut-off value, ≥31.91 IU/mL), whereas the Abbott Alinity IgG assay detected all 334 samples as positive (above the cut-off value ≥ 7.1 BAU/mL).

This showed high agreement between both detection systems. (Supplementarys. Block E: Crosstab approach, Tables 32,22).

In contrast to many professions that transitioned to remote work during the pandemic, healthcare workers remained continuously exposed to potential sources of SARS-CoV-2 transmission (3). This was particularly relevant in dental settings, where aerosol-generating procedures and close patient contact increased occupational exposure risks. Consequently, reliable and rapidly deployable methods for assessing vaccine-induced immunity became especially relevant for occupational health surveillance in these populations. Similar observations have been reported in studies of dental professionals, a population characterized by sustained occupational exposure to aerosol-generating procedures. Duś-Ilnicka (21) et al. demonstrated significant post-vaccination increases in both SARS-CoV-2 IgG concentrations and neutralizing antibody responses using complementary serological approaches. Their findings support the concept that quantitative IgG assays, such as the Abbott Alinity system, and assays reflecting neutralizing antibody activity provide related but not identical information regarding vaccine-induced humoral immunity. In this context, the observed agreement between Abbott and EDELVITAL measurements in the present study supports the potential use of fluorescence immunography as a practical adjunct to established laboratory-based serological methods, particularly in occupational health settings. The results of this study show that the rapid point-of-care device has comparable diagnostic performance to the established Abbott Alinity I CMIA assay but offers practical added value due to its speed and applicability. Therefore, this point-of-care device is a potentially valuable tool for the rapid assessment of the humoral vaccine response, especially in situations where timely decisions on booster vaccinations are required.

Age- and sex-related effects

Age and sex have been reported as potential modifiers of SARS-CoV-2 antibody responses (26, 27); however, the magnitude and consistency of these effects vary across studies and settings. In another large longitudinal cohort after two-dose BNT162b2 vaccination, humoral responses were lower in men and in older individuals (≥65 years), indicating that demographic factors can contribute to inter-individual variability (12). In contrast, in infection-derived immunity, antibody magnitude appears to be strongly shaped by clinical phenotype, with more robust antibody responses reported in individuals experiencing more severe COVID-19 symptoms early after infection (13). Our data indicate that age and sex had only a limited effect on antibody responses. This might be related to the fact that, as already reported, there is no overall difference in SARS-CoV-2 antibody responses in between age groups (30). Sex-related effects were largely negligible, with comparable IgG and nAb levels between male and female participants. Apart from the lower pre-booster nAb in men, this isolated finding is consistent with reports of stronger humoral responses in females after the second vaccination (12). Taken together, these data suggest that demographic factors may modulate antibody levels, but infection severity (where applicable), immune status, and time since antigen exposure can represent major determinants of measured humoral responses.

Booster effect and quantitative comparison

The observed positive correlations between IgG levels and neutralizing antibodies prior to booster vaccination are consistent with previous reports demonstrating an association between binding and functional antibody responses following SARS-CoV-2 vaccination. Levine et al. reported a strong correlation between IgG and neutralizing antibody titers for up to six months after the second vaccine dose (Spearman’s ρ ranging from 0.68 to 0.75), although the relationship between the two parameters varied over time, likely reflecting different kinetics of binding and functional antibodies (12). In the present study, a strong correlation between IgG and neutralizing antibodies prior to the third dose was also observed (Spearman’s rs = 0.631, p < 0.001), while additional moderate correlations were detected between pre-booster nAbs and post-booster IgG levels (rs = 0.528, p < 0.001) as well as between IgG levels before and after the booster (rs = 0.382, p = 0.012). Together, these findings support the general association between IgG responses and neutralizing capacity while also suggesting variability depending on sampling time points and cohort characteristics (12).

Nouanesengsy et al., who investigated individuals with prior natural SARS-CoV-2 infection followed by vaccination, observed a marked increase in both IgG and neutralizing antibody responses after the first vaccine dose, whereas no additional increase was detected after the second dose (13). This pattern might be attributed to immune priming induced by prior infection and matched our observations after the third booster. Although the present study focused on booster vaccination rather than hybrid immunity, the observed association between pre-booster neutralizing antibodies and post-booster IgG levels may similarly reflect the influence of pre-existing immune responses on subsequent antibody production. The pronounced increase observed in the paired-sample analysis likely reflects the true immunological effect of booster vaccination, as longitudinal comparisons within the same individuals reduce variability caused by heterogeneous baseline antibody levels. Despite these associations, the comparability between antibody responses measured before and after the third vaccination remained limited. Previous studies have demonstrated that SARS-CoV-2 antibody responses are highly heterogeneous between individuals, with antibody titers spanning a wide range across populations. In addition, the interpretation of post-booster comparisons in our cohort is limited by the smaller number of samples available after the third vaccination. Furthermore, binding IgG assays and neutralization assays measure different immunological aspects of the humoral response. While binding assays quantify antibody recognition of viral antigens, neutralization assays assess the functional ability of antibodies to inhibit viral entry. Consequently, high IgG titers do not necessarily correspond to strong neutralizing activity. Such differences in assay principles and antibody kinetics may partially explain the moderate correlations observed in the present study (10, 13). Previous studies have also suggested that antibody responses may reach a temporary plateau following repeated antigen exposure, particularly after the second vaccine dose (31) or previous antigen contact (32). Regression modeling has suggested that neutralizing antibody titers may decline more rapidly, with an estimated half-life of approximately 14.7 weeks compared with 31.4 weeks for IgG levels (28). Similarly, Vanyshalla et al. reported that not all vaccinated individuals develop measurable neutralizing antibody responses following SARS-CoV-2 infection despite detectable IgG antibodies.

Diagnostic performance (crosstab analysis)

The primary aim of this study was to evaluate the suitability of the EDELVITAL Fluorescence Immunochromatography Test system for determining a sufficient antibody response using a binary classification (positive/negative). The overall agreement between the rapid test system and the laboratory-based Abbott Alinity I SARS-CoV-2 IgG II Quant assay was high, with concordant positive results in 95.5% of samples (Table 3). The results indicate a high level of agreement between both assays in identifying vaccine-induced humoral immune responses, despite targeting different components of the antibody response.

The rapid assay demonstrated a high level of concordance with the established laboratory-based platform while providing information on neutralizing antibody activity in a point-of-care setting. Only 15 samples (4.5%) showed discordant results, characterized by detectable IgG levels but negative neutralizing antibody responses. Such discrepancies are biologically plausible, due to the antibody kinetics as discussed above. Several methodological aspects should be additionally considered when interpreting the concordance of diagnostic performance. Agreement between assays does not necessarily imply equivalence in analytical performance because both methods assess different aspects of the humoral immune response. The Abbott assay quantifies anti-spike IgG antibodies, whereas the EDELVITAL assay evaluates neutralizing antibody activity. The Abbott assay provides quantitative measurements of anti-spike IgG antibodies, whereas the EDELVITAL test produces a qualitative binary output. The rapid assay cannot capture differences in antibody concentrations, particularly near diagnostic thresholds or in cases of declining antibody titers. High concordance primarily indicates that the rapid test identifies samples with clearly detectable antibody levels rather than demonstrating equivalent quantitative performance. The interpretation of serological results remains inherently limited by the nature of antibody detection. Rapid serological tests detect binding antibodies, which reflect previous exposure or vaccination but do not directly measure functional neutralizing capacity, as highlighted in an analytical review of rapid diagnostic technologies (25). Similarly, the relationship between binding antibodies and protective immunity is complex. A study on SARS-CoV-2 serological assays emphasizes that binding antibody assays do not necessarily reflect neutralizing activity, although correlations between the two have been reported (24). In our cohort, moderate-to-strong correlations between IgG levels and neutralizing antibodies were observed, suggesting that IgG measurements can provide a reasonable approximation of neutralizing capacity at the population level.

Neutralizing antibodies have been proposed as an important correlate of protection against SARS-CoV-2 infection (33), with several studies demonstrating quantitative relationships between neutralization titers and the risk of symptomatic infection. Although clinical protection was not assessed in the present study, the detection of neutralizing antibodies in the participants suggests the presence of measurable humoral immunity in this vaccinated cohort. However, due to its binary output and the inherent limitations of binding antibody detection, the assay cannot replace quantitative laboratory immunoassays or functional neutralization tests when detailed characterization of the immune response is required.

In this context, EDELVITAL should be viewed as a complementary tool rather than a replacement for established laboratory-based serological assays. While laboratory-based assays remain the gold standard for the detailed characterization of antibody functionality, rapid point-of-care tests offer advantages in terms of simplicity, rapid turnaround time, and logistical feasibility. These characteristics may facilitate their use in real-world settings, such as occupational health programs or large-scale screening strategies, where rapid assessment of antibody responses could support individualized vaccination decisions.

Limitations

This study examined two vaccination regimens: (1) two doses of BNT162b2 and (2) the same primary series followed by an mRNA-1273 booster dose. Because only limited numbers of participants had received other vaccine types or combinations, we were unable to compare immune responses or effectiveness across different vaccination platforms.

Furthermore, this study focused on comparing IgG and neutralizing antibody responses in vaccinated individuals and did not verify negative serological statuses in the unvaccinated population. The interpretation of EDELVITAL measurements should also be considered in the context of SARS-CoV-2 variant evolution. The assay was developed using an RBD antigen derived from the ancestral Wuhan strain, which was the standard reference antigen when the study was initiated. During the study period, Omicron variant and its sublineages became increasingly dominant. Antigenic differences between ancestral and Omicron-derived strains, variant-specific antibody recognition may have been affected. This could partially explain discordant findings in which detectable anti-SARS-CoV-2 IgG concentrations were not accompanied by corresponding EDELVITAL measurements. Therefore, the results should be interpreted within the temporal and epidemiological context in which the study was conducted. Owing to the continuous emergence of new SARS-CoV-2 variants, it was not possible to assess variant-specific immune responses. A direct comparison with live-virus or microneutralization assays was not performed in the present study and therefore conclusions regarding true neutralizing capacity remain limited. Previous studies have demonstrated that neutralizing antibody responses may differ substantially between SARS-CoV-2 variants despite comparable levels of vaccine-induced antibodies. In particular, Omicron-lineage variants show markedly reduced susceptibility to neutralization compared with earlier strains (36), reflecting the impact of accumulated spike protein mutations on antibody recognition. Furthermore, considerable interindividual variability in neutralizing responses has been reported even among similarly vaccinated individuals. These observations may partly explain the moderate correlations observed between quantitative IgG measurements and EDELVITAL results, as well as cases in which detectable anti-SARS-CoV-2 IgG concentrations were not accompanied by corresponding EDELVITAL measurements. Future studies directly comparing EDELVITAL measurements with classical microneutralization assays across different SARS-CoV-2 variants would be valuable to further define assay sensitivity and clinical relevance. Such analyses would be of considerable value for future pandemic waves or potential epidemics. Interpretation of SARS-CoV-2 serological findings remains challenging, particularly in occupationally exposed healthcare populations. Although participants were routinely monitored by institutional surveillance programs and were excluded from the present study when tested positive for SARS-CoV.2, previous asymptomatic or otherwise undetected infections cannot be completely excluded. Consequently, measured antibody responses may reflect a combination of vaccine-induced immunity and prior natural exposure. Similar challenges have been reported in studies of healthcare and dental professionals, highlighting the complexity of distinguishing infection-induced from vaccine-induced antibody responses based solely on serological measurements (21, 28,36).Therefore, antibody concentrations and surrogate neutralizing antibody measurements should be interpreted within the broader clinical and epidemiological context. A major limitation of the present study is the absence of a direct neutralization assay to verify the functional neutralizing capacity of the detected antibodies, and this clearly limits the ability to formally validate the EDELVITAL Fluorescence Immunochromatography Test system. Consequently, the conclusions drawn are restricted to assay performance rather than a definitive assessment of protective neutralizing activity, underscoring the need for future studies incorporating classical neutralization assays.

This study demonstrates the high degree of analytical agreement between the automated Abbott Alinity-I IgG II Quant Assay and the EDELVITAL fluorescence immunochromatography test for determining humoral immune responses to SARS-CoV-2 vaccination.

This underscores that standardized rapid tests are an increasingly valid alternative to laboratory-based methods when they are calibrated to the WHO reference standard 20/136, as is the case here.

Nevertheless, the use of such tests in clinical decision-making should always be considered in the context of other immunological parameters, and the individual patient’s situation should always be taken into account. Overall, the results show that the combination of precise laboratory analysis and practical rapid test technology can contribute to more efficient vaccine response monitoring. The EDELVITAL test proved to be a reliable and practical tool for the supplementary assessment of neutralizing antibody activity and can thus bridge the gap between the laboratory and everyday clinical practice and protection in health workers. From an occupational health perspective, rapid and easily deployable serological assays may support the longitudinal monitoring of vaccine-induced immune responses in healthcare workers and other populations at increased risk of exposure. Although laboratory-based methods remain essential for comprehensive antibody characterization, fluorescence immunography offers the advantage of rapid result generation and minimal infrastructure requirements. Consequently, such assays may represent a useful adjunct for repeated immune-status assessments in occupational health programs and large-scale surveillance settings.