Imaging-based characterization of tumoral heterogeneity for personalized cancer treatment.

With personalized tumor therapy, understanding and addressing the heterogeneity of malignant tumors is becoming increasingly important. Heterogeneity can be found within one lesion (intralesional) and between several tumor lesions emerging from one primary tumor (interlesional). The heterogeneous tumor cells may show a different response to treatment due to their biology, which in turn influences the outcome of the affected patients and the choice of therapeutic agents. Therefore, both intra- and interlesional heterogeneity should be addressed at the diagnostic stage. While genetic and biological heterogeneity are important parameters in molecular tumor characterization and in histopathology, they are not yet addressed routinely in medical imaging. This article summarizes the recently established markers for tumor heterogeneity in imaging as well as heterogeneous/mixed response to therapy. Furthermore, a look at emerging markers is given. The ultimate goal of this overview is to provide comprehensive understanding of tumor heterogeneity and its implications for radiology and for communication with interdisciplinary teams in oncology. KEY POINTS:: · Tumor heterogeneity can be described within one lesion (intralesional) or between several lesions (interlesional).. · The heterogeneous biology of tumor cells can lead to a mixed therapeutic response and should be addressed in diagnostics and the therapeutic regime.. · Quantitative image diagnostics can be enhanced using AI, improved histopathological methods, and liquid profiling in the future..

Triptychon: Usability evaluation and implementation of a web-based application for patients' lab and vital parameters.

Background: A major challenge in healthcare is the interpretation of the constantly increasing amount of clinical data of interest to inpatients for diagnosis and therapy. It is vital to accurately structure and represent data from different sources to help clinicians make informed decisions. Objective: We evaluated the usability of our tool 'Triptychon' - a three-part visualisation dashboard of essential patients' medical data provided by a direct overview of their hospitalisation information, laboratory, and vital parameters over time. Methods: The study followed a cohort of 20 participants using the mixed-methods approach, including interviews and the usability questionnaires, Health Information Technology Usability Evaluation Scale (Health-ITUES), and User Experience Questionnaire (UEQ). The participant's interactions with the dashboard were also observed. A thematic analysis approach was applied to analyse qualitative data and the quantitative data's task completion time and success rates. Results: The usability evaluation of the visualisation dashboard revealed issues relating to the terminology used in the user interface and colour coding in its left and middle panels. The Health-ITUES score was 3.72 (standard deviation (SD) = 1.0), and the UEQ score was 1.6 (SD = 0.74). The study demonstrated improvements in intuitive dashboard use and overall satisfaction with using the dashboard daily. Conclusion: The Triptychon dashboard is a promising new tool for medical data presentation. We identified design and layout issues of the dashboard for improving its usability in routine clinical practice. According to users' feedback, the three panels on the dashboard provided a holistic view of a patient's hospital stay.

Exploring the Synergistic Potential of Radiomics and Laboratory Biomarkers for Enhanced Identification of Vulnerable COVID-19 Patients.

BACKGROUND: Severe courses and high hospitalization rates were ubiquitous during the first pandemic SARS-CoV-2 waves. Thus, we aimed to examine whether integrative diagnostics may aid in identifying vulnerable patients using crucial data and materials obtained from COVID-19 patients hospitalized between 2020 and 2021 (n = 52). Accordingly, we investigated the potential of laboratory biomarkers, specifically the dynamic cell decay marker cell-free DNA and radiomics features extracted from chest CT. METHODS: Separate forward and backward feature selection was conducted for linear regression with the Intensive-Care-Unit (ICU) period as the initial target. Three-fold cross-validation was performed, and collinear parameters were reduced. The model was adapted to a logistic regression approach and verified in a validation naïve subset to avoid overfitting. RESULTS: The adapted integrated model classifying patients into "ICU/no ICU demand" comprises six radiomics and seven laboratory biomarkers. The models' accuracy was 0.54 for radiomics, 0.47 for cfDNA, 0.74 for routine laboratory, and 0.87 for the combined model with an AUC of 0.91. CONCLUSION: The combined model performed superior to the individual models. Thus, integrating radiomics and laboratory data shows synergistic potential to aid clinic decision-making in COVID-19 patients. Under the need for evaluation in larger cohorts, including patients with other SARS-CoV-2 variants, the identified parameters might contribute to the triage of COVID-19 patients.

Results from an IFCC global survey on laboratory practices for the analysis of circulating tumor DNA.

BACKGROUND: The clinical validity of ctDNA analysis as a diagnostic, prognostic and predictive biomarker has been demonstrated in many studies. The rapid spread of tests for the analysis of ctDNA raises questions regarding their standardization and quality assurance. The aim of this study was to provide a global overview of the test methods, laboratory procedures and quality assessment practices using ctDNA diagnostics. METHODS: The Molecular Diagnostics Committee of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC C-MD) conducted a survey among international laboratories performing ctDNA analysis. Questions on analytical techniques, test parameters, quality assurance and the reporting of findings were included. RESULTS: A total of 58 laboratories participated in the survey. The majority of the participating laboratories (87.7 %) performed testing for patient care. Most laboratories conducted their assays for lung cancer (71.9 %), followed by colorectal (52.6 %) and breast (40.4 %) cancer, and 55.4 % of the labs used ctDNA analysis for follow-up/monitoring of treatment-resistant alterations. The most frequent gene analysed was EGFR (75.8 %), followed by KRAS (65.5 %) and BRAF (56.9 %). Participation in external quality assessment programs was reported by only 45.6 % of laboratories. CONCLUSIONS: The survey indicates that molecular diagnostic methods for the analysis of ctDNA are not standardized across countries and laboratories. Furthermore, it reveals a number of differences regarding sample preparation, processing and reporting test results. Our findings indicate that ctDNA testing is being conducted without sufficient attention to analytical performance between laboratories and highlights the need for standarisation of ctDNA analysis and reporting in patient care.

Heterologous Vector-mRNA Based SARS-CoV-2 Vaccination Strategy Appears Superior to a Homologous Vector-Based Vaccination Scheme in German Healthcare Workers Regarding Humoral SARS-CoV-2 Response Indicating a High Boosting Effect by mRNA Vaccines.

BACKGROUND: Longitudinal humoral SARS-CoV-2 (severe acute respiratory syndrome coronavirus type 2) immunity for up to 15 months due to vaccination, the efficacy of vaccination strategies (homologous, vector-vector versus heterologous, vector-mRNA), the influence of vaccination side effects, and the infection rate in German healthcare workers need to be investigated. METHODS: In this study, 103 individuals vaccinated against SARS-CoV-2 were enrolled to examine their anti-SARS-CoV-2 anti-N- and anti-RBD/S1-Ig levels. A total of 415 blood samples in lithium heparin tubes were prospectively obtained, and a structured survey regarding medical history, type of vaccine, and vaccination reactions was conducted. RESULTS: All participants demonstrated a humoral immune response, among whom no values decreased below the positivity cutoff. Five to six months after the third vaccination, three participants showed anti-RBD/S1 antibodies of less than 1000 U/mL. We observed higher levels for heterologous mRNA-/vector-based combinations compared to pure vector-based vaccination after the second vaccination, which is harmonized after a third vaccination with the mRNA-vaccine only in both cohorts. The incidence of vaccine breakthrough in a highly exposed cohort was 60.3%. CONCLUSION: Sustained long-term humoral immunity was observed, indicating the superiority of a heterologous mRNA-/vector-based combination compared to pure vector-based vaccination. There was longevity of anti-RBD/S1 antibodies of at least 4 and up to 7 months without external stimulus. Regarding vaccination reactogenity, the occurrence of local symptoms as pain at the injection site was increased after the first mRNA application compared to the vector-vector cohort with a general decrease in adverse events at later vaccination time points. Overall, a correlation between the humoral vaccination response and vaccination side effects was not observed. Despite the high prevalence of vaccine breakthroughs, these only occurred in the later course of the study when more infectious variants, which are, however, associated with milder courses, were present. These results provide insights into vaccine-related serologic responses, and the study should be expanded using additional vaccine doses and novel variants in the future.

Humoral SARS-CoV-2 Immune Response in COVID-19 Recovered Vaccinated and Unvaccinated Individuals Related to Post-COVID-Syndrome.

BACKGROUND: The duration of anti-SARS-CoV-2-antibody detectability up to 12 months was examined in individuals after either single convalescence or convalescence and vaccination. Moreover, variables that might influence an anti-RBD/S1 antibody decline and the existence of a post-COVID-syndrome (PCS) were addressed. METHODS: Forty-nine SARS-CoV-2-qRT-PCR-confirmed participants completed a 12-month examination of anti-SARS-CoV-2-antibody levels and PCS-associated long-term sequelae. Overall, 324 samples were collected. Cell-free DNA (cfDNA) was isolated and quantified from EDTA-plasma. As cfDNA is released into the bloodstream from dying cells, it might provide information on organ damage in the late recovery of COIVD-19. Therefore, we evaluated cfDNA concentrations as a biomarker for a PCS. In the context of antibody dynamics, a random forest-based logistic regression with antibody decline as the target was performed and internally validated. RESULTS: The mean percentage dynamic related to the maximum measured value was 96 (±38)% for anti-RBD/S1 antibodies and 30 (±26)% for anti-N antibodies. Anti-RBD/S1 antibodies decreased in 37%, whereas anti-SARS-CoV-2-anti-N antibodies decreased in 86% of the subjects. Clinical anti-RBD/S1 antibody decline prediction models, including vascular and other diseases, were cross-validated (highest AUC 0.74). Long-term follow-up revealed no significant reduction in PCS prevalence but an increase in cognitive impairment, with no indication for cfDNA as a marker for a PCS. CONCLUSION: Long-term anti-RBD/S1-antibody positivity was confirmed, and clinical parameters associated with declining titers were presented. A fulminant decrease in anti-SARS-CoV-2-anti-N antibodies was observed (mean change to maximum value 30 (±26)%). Anti-RBD/S1 antibody titers of SARS-CoV-2 recovered subjects boosted with a vaccine exceeded the maximum values measured after single infection by 235 ± 382-fold, with no influence on preexisting PCS. PCS long-term prevalence was 38.6%, with an increase in cognitive impairment compromising the quality of life. Quantified cfDNA measured in the early post-COVID-19 phase might not be an effective marker for PCS identification.

Results of a European-Wide External Quality Assessment (EQA) Scheme for Serological Detection of Anti-SARS-CoV-2 (CoVimm)-Pitfalls of Routine Application.

BACKGROUND: During the last two years, a variety of assays for the serological detection of antibodies to the new SARS-CoV-2 virus have been launched and used as part of standard care in many laboratories. The pace with which these tests have been introduced into routine care emphasizes the importance of quality measures for analytical methods, particularly with regard to the implications of results for clinical and epidemiologic decisions. Accuracy, reliability and comparability of analytical test results are thus essential, and here external quality assessment (EQA) is the most important quality assurance tool. It allows us to achieve harmonization of test methods as a prerequisite for a high standard of performance for laboratory and analytical techniques and their interpretation. METHODS: This EQA scheme consisted of pre-characterized clinical biospecimens dedicated to the analysis of anti-SARS-CoV-2 IgG total antibodies and differentiation into spike protein-specific IgG antibodies against SARS-CoV-2 (anti-S-SARS-CoV-2) and nucleocapsid-specific IgG antibodies against SARS-CoV-2 (anti-N-SARS-CoV-2). RESULTS: A total of 239 laboratories across Europe participated in this scheme, called CoVimm. In detail, 536 results for anti-SARS-CoV-2 IgG, 431 results for anti-S-SARS-CoV-2 IgG, and 200 results for anti-N-SARS-CoV-2 IgG were reported. Based on the pre-defined thresholds, the success rates for the determination of anti-S-SARS-CoV-2 IgG and anti-N-SARS-CoV-2 IgG were 96% and 90%, respectively. Interestingly, only 64% of the participating laboratories successfully passed the EQA scheme for the determination of total anti-SARS-CoV-2 IgG. CONCLUSIONS: This EQA revealed serious concerns regarding the reliability and appropriate use of anti-SARS-CoV-2 antibody assays in routine care. In addition to the wide heterogeneity of different assays used by participating laboratories, a lack of standardization and harmonization is also evident. This is of particular importance for reliable and clinically meaningful interpretation of test results.

The Potential and Emerging Role of Quantitative Imaging Biomarkers for Cancer Characterization.

Similar to the transformation towards personalized oncology treatment, emerging techniques for evaluating oncologic imaging are fostering a transition from traditional response assessment towards more comprehensive cancer characterization via imaging. This development can be seen as key to the achievement of truly personalized and optimized cancer diagnosis and treatment. This review gives a methodological introduction for clinicians interested in the potential of quantitative imaging biomarkers, treating of radiomics models, texture visualization, convolutional neural networks and automated segmentation, in particular. Based on an introduction to these methods, clinical evidence for the corresponding imaging biomarkers-(i) dignity and etiology assessment; (ii) tumoral heterogeneity; (iii) aggressiveness and response; and (iv) targeting for biopsy and therapy-is summarized. Further requirements for the clinical implementation of these imaging biomarkers and the synergistic potential of personalized molecular cancer diagnostics and liquid profiling are discussed.

Anosmia Testing as Early Detection of SARS-CoV-2 Positivity; A Prospective Study under Screening Conditions.

Sudden onset of anosmia is a phenomenon often associated with developing COVID-19 disease and has even been described as an initial isolated symptom in individual cases. In this case-control study, we investigated the feasibility of this condition as a suitable screening test in a population at risk. We performed a prospective study with a total of 313 subjects with suspected SARS-CoV-2 infection. In parallel to routine PCR analysis, a modified commercial scent test was performed to objectify the presence of potential anosmia as a predictor of SARS-CoV-2 positivity. Furthermore, a structured interview assessment of the participants was conducted. A total of 12.1% of the study participants had molecular genetic detection of SARS-CoV-2 infection in the nasopharyngeal swab. It could be demonstrated that these subjects had a significantly weaker olfactory identification performance of the scents. Further analysis of the collected data from the scent test and medical history via random forest (Boruta) algorithm showed that no improvement of the prediction power was achieved by this design. The assay investigated in this study may be suitable for screening general olfactory function. For the screening of COVID-19, it seems to be affected by too many external and internal biases and requires too elaborate and selective pre-test screening.

Integrated diagnostics.

BACKGROUND: Integrated diagnostics is increasingly gaining scientific traction as it promises to address several challenges currently facing diagnostic medicine. These challenges range from the need for improved diagnostic accuracy to optimized timing of diagnostic procedures, to the variety of diagnostic markers and thus the complexity of their interpretation, and finally to economic pressure. METHODICAL INNOVATIONS: While many of these challenges may be difficult to solve with a monomodal approach, the integration of laboratory markers and imaging procedures promises to allow both disciplines to achieve their actual clinical potential. Combining complementary diagnostic approaches can help to improve the interpretation of measurements, provide a better cost-effectiveness particularly when cutting-edge techniques are used for specific indications, and facilitate optimized timing and rational choice of appropriate diagnostic approaches for disease surveillance. Furthermore, close interdisciplinary assessment of diagnostic results will increase diagnostic accuracy and will enable selection of specific patient cohorts at increased risk for certain diseases who are suitable for further testing. CONCLUSION: The potential of an integrated diagnostic approach represents a strategic goal for diagnostic disciplines as it achieves better visibility and greater clinical impact. In addition to close collaboration among relevant diagnostic experts, an appropriate structure for integrated data evaluation needs to be established to provide actionable health guidance so that integrated diagnostics can be implemented in standard care.

Identification of CT Imaging Phenotypes of Colorectal Liver Metastases from Radiomics Signatures-Towards Assessment of Interlesional Tumor Heterogeneity.

(1) Background: Tumoral heterogeneity (TH) is a major challenge in the treatment of metastatic colorectal cancer (mCRC) and is associated with inferior response. Therefore, the identification of TH would be beneficial for treatment planning. TH can be assessed by identifying genetic alterations. In this work, a radiomics-based approach for assessment of TH in colorectal liver metastases (CRLM) in CT scans is demonstrated. (2) Methods: In this retrospective study, CRLM of mCRC were segmented and radiomics features extracted using pyradiomics. Unsupervised k-means clustering was applied to features and lesions. Feature redundancy was evaluated by principal component analysis and reduced by Pearson correlation coefficient cutoff. Feature selection was conducted by LASSO regression and visual analysis of the clusters by radiologists. (3) Results: A total of 47 patients' (36% female, median age 64) CTs with 261 lesions were included. Five clusters were identified, and the categories small disseminated (n = 31), heterogeneous (n = 105), homogeneous (n = 64), mixed (n = 59), and very large type (n = 2) were assigned based on visual characteristics. Further statistical analysis showed correlation (p < 0.01) of clusters with sex, primary location, T- and N-status, and mutational status. Feature reduction and selection resulted in the identification of four features as a final set for cluster definition. (4) Conclusions: Radiomics features can characterize TH in liver metastases of mCRC in CT scans, and may be suitable for a better pretherapeutic classification of liver lesion phenotypes.

Liquid Profiling for Cancer Patient Stratification in Precision Medicine-Current Status and Challenges for Successful Implementation in Standard Care.

Circulating tumor DNA (ctDNA), accurately described by the term liquid profiling (LP), enables real-time assessment of the tumor mutational profile as a minimally invasive test and has therefore rapidly gained traction, particular for the management of cancer patients. By LP, tumor-specific genetic alterations can be determined as part of companion diagnostics to guide selection of appropriate targeted therapeutics. Because LP facilitates longitudinal monitoring of cancer patients, it can be used to detect acquired resistant mechanisms or as a personalized biomarker for earlier detection of disease recurrence, among other applications. However, LP is not yet integrated into routine care to the extent that might be expected. This is due to the lack of harmonization and standardization of preanalytical and analytical workflows, the lack of proper quality controls, limited evidence of its clinical utility, heterogeneous study results, the uncertainty of clinicians regarding the value and appropriate indications for LP and its interpretation, and finally, the lack of reimbursement for most LP tests. In this review, the value proposition of LP for cancer patient management and treatment optimization, the current status of implementation in standard care, and the main challenges that need to be overcome are discussed in detail.

The value proposition of integrative diagnostics for (early) detection of cancer. On behalf of the EFLM interdisciplinary Task and Finish Group "CNAPS/CTC for early detection of cancer".

Disruptive imaging and laboratory technologies can improve clinical decision processes and outcomes in oncology. However, certain obstacles must be overcome before these technologies can be fully implemented as part of the standard for care. An integrative diagnostic approach represents a unique opportunity to unleash the full diagnostic potential and paves the way towards personalized cancer diagnostics. To meet this demand, an interdisciplinary Task Force of the EFLM was initiated as a consequence of an EFLM/ESR during the CELME 2019 meeting in order to evaluate the clinical value of CNAPS/CTC (circulating nucleic acids in plasma and serum/circulating tumor cells) in early detection of cancer. Here, an overview of current disruptive techniques, their clinical implications and potential value of an integrative diagnostic approach is provided. Furthermore, requirements such as the establishment of diagnostic tumor boards, development of adequate software solutions and a change of mindset towards a new generation of diagnosticians providing actionable health information are presented. This development has the potential to elevate the position and clinical recognition of diagnosticians.

Liquid profiling of circulating tumor DNA in colorectal cancer: steps needed to achieve its full clinical value as standard care.

The analysis of circulating tumor DNA (ctDNA) is at the threshold of implementation into standard care for colorectal cancer (CRC) patients. However, data about the clinical utility of liquid profiling (LP), its acceptance by clinicians, and its integration into clinical workflows in real-world settings remain limited. Here, LP tests requested as part of routine care since 2016 were retrospectively evaluated. Results show restrained request behavior that improved moderately over time, as well as reliable diagnostic performance comparable to translational studies, with an overall agreement of 91.7%. Extremely low ctDNA levels at < 0.1% in over 20% of cases, a high frequency of concomitant driver mutations (in up to 14% of cases), and ctDNA levels reflecting the clinical course of disease were revealed. However, certain limitations hampering successful translation of ctDNA into clinical practice were uncovered, including the lack of clinically relevant ctDNA thresholds, appropriate time points of LP requests, and integrative evaluation of ctDNA, imaging, and clinical findings. In conclusion, these results highlight the potential clinical value of LP for CRC patient management and demonstrate issues that need to be addressed for successful long-term implementation in clinical workflows.

Economic assessment of molecular imaging in the oncology treatment process.

The development towards targeted treatments in oncology has been accompanied by significant improvements in molecular imaging. Yet, broad application of novel imaging techniques has partly been slowed down due to economical considerations. Building on the broad positive evidence of its diagnostic accuracy, modelling of effects on long-term costs and effectiveness may help to foster a broader application and acceptance of comprehensive molecular imaging techniques, such as PET/MRI. In this article, common economic evaluation techniques and cost-effectiveness analysis (CEA) evaluation methods will be introduced including Markov models and incremental cost-effectiveness ratios (ICER). This is complemented with a review of literature on recently published cost-effectiveness of molecular imaging. Additionally, the strategic relevance of CEAs for the molecular imaging community is discussed and combined with a global outlook.

Recruitment of highly cytotoxic CD8+ T cell receptors in mild SARS-CoV-2 infection.

T cell immunity is crucial for control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and has been studied widely on a quantitative level. However, the quality of responses, in particular of CD8+ T cells, has only been investigated marginally so far. Here, we isolate T cell receptor (TCR) repertoires specific for immunodominant SARS-CoV-2 epitopes restricted to common human Leukocyte antigen (HLA) class I molecules in convalescent individuals. SARS-CoV-2-specific CD8+ T cells are detected up to 12 months after infection. TCR repertoires are diverse, with heterogeneous functional avidity and cytotoxicity toward virus-infected cells, as demonstrated for TCR-engineered T cells. High TCR functionality correlates with gene signatures that, remarkably, could be retrieved for each epitope:HLA combination analyzed. Overall, our data demonstrate that polyclonal and highly functional CD8+ TCRs-classic features of protective immunity-are recruited upon mild SARS-CoV-2 infection, providing tools to assess the quality of and potentially restore functional CD8+ T cell immunity.

Preclinical evaluation of eltrombopag in a PDX model of myelodysplastic syndromes.

Preclinical research of myelodysplastic syndromes (MDSs) is hampered by a lack of feasible disease models. Previously, we have established a robust patient-derived xenograft (PDX) model for MDS. Here we demonstrate for the first time that this model is applicable as a preclinical platform to address pending clinical questions by interrogating the efficacy and safety of the thrombopoietin receptor agonist eltrombopag. Our preclinical study included n = 49 xenografts generated from n = 9 MDS patient samples. Substance efficacy was evidenced by FACS-based human platelet quantification and clonal bone marrow evolution was reconstructed by serial whole-exome sequencing of the PDX samples. In contrast to clinical trials in humans, this experimental setup allowed vehicle- and replicate-controlled analyses on a patient-individual level deciphering substance-specific effects from natural disease progression. We found that eltrombopag effectively stimulated thrombopoiesis in MDS PDX without adversely affecting the patients' clonal composition. In conclusion, our MDS PDX model is a useful tool for testing new therapeutic concepts in MDS preceding clinical trials.

SARS-CoV-2 Antibody Testing - How to Increase the Positive Predictive Value.

BACKGROUND: Despite increasing COVID-19 infection rates, low overall prevalence resulting in a poor positive predictive value (PPV) of serological tests requires strategies to increase specificity. We therefore investigated a dual diagnostic strategy and evaluated the correlation between the severity of a SARS-CoV-2 infection and the detectable immune-response. METHODS: Participants were systematically categorized into positive and control cohorts and a probability score of COVID-19 was calculated based on clinical symptoms. Six hundred eighty-two serum samples were analyzed using a highly specific high-throughput system. Combining the serological test result and probability score was performed as a dual diagnostic strategy. RESULTS: Specificity of 99.61% and sensitivity of 86.0% were the basis of our approach. A dual diagnostic strategy led to increased pre-test probability and thus to a test specificity of 100%. In a flu-like symptomatic population, we estimated a COVID-prevalence of 4.79%. Moreover, we detected significantly higher antibody values in patients with fever than without fever. CONCLUSIONS: Based on sensitivity and specificity results of our study being in line with previous findings, we demonstrated a dual assessment strategy including a symptom-based probability score and serological testing to increase the PPV. Moreover, the presence of fever seems to trigger a stronger immune-response.

Assessing the Quality of Serological Testing in the COVID-19 Pandemic: Results of a European External Quality Assessment (EQA) Scheme for Anti-SARS-CoV-2 Antibody Detection.

External quality assessment (EQA) is a key instrument for achieving harmonization, and thus a high quality, of diagnostic procedures. As reliable test results are crucial for accurate assessment of SARS-CoV-2 infection prevalence, vaccine response, and immunity, and thus for successful management of the ongoing COVID-19 pandemic, the Reference Institute for Bioanalytics (RfB) was the first EQA provider to offer an open scheme for anti-SARS-CoV-2 antibody detection. The main objectives of this EQA were (i) to gain insights into the current diagnostic landscape and the performance of serological tests in Europe and (ii) to provide recommendations for diagnostic improvements. Within the EQA, a blinded panel of precharacterized human serum samples with variable anti-SARS-CoV-2 antibody titers was provided for detection of anti-SARS-CoV-2 IgG, IgA, and IgM antibodies. Across the three distribution rounds in 2020, 284 laboratories from 22 countries reported a total of 3,744 results for anti-SARS-CoV-2 antibody detection using more than 24 different assays for IgG. Overall, 97/3,004 results were false for anti-SARS-CoV-2 IgG, 88/248 for IgA, and 34/124 for IgM. Regarding diagnostic sensitivity and specificity, substantial differences were found between the different assays used, as well as between certified and noncertified tests. For cutoff samples, a drop in the diagnostic sensitivity to 46.3% and high interlaboratory variability were observed. In general, this EQA highlights the current variability of anti-SARS-CoV-2 antibody detection, technical limitations with respect to cutoff samples, and the lack of harmonization of testing procedures. Recommendations are provided to help laboratories and manufacturers further improve the quality of anti-SARS-CoV-2 serological diagnostics.