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Investigation of Antibody Pharmacokinetics in Male Reproductive System and Its Characterization Using a Translational PBPK Model
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Clinical Scaleup of Humanized AnnA1 Antibody Yielded Unexpected High Reticuloendothelial (RES) Uptake in Mice
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Immune Cell Engagers: Advancing Precision Immunotherapy for Cancer Treatment
Journal Description
Antibodies
Antibodies
is an international, peer-reviewed, open access journal on immunoglobulins, published quarterly online by MDPI.
- Open Access— free for readers, with article processing charges (APC) paid by authors or their institutions.
- High Visibility: indexed within Scopus, ESCI (Web of Science), PubMed, PMC, Embase, CAPlus / SciFinder, and other databases.
- Journal Rank: CiteScore - Q2 (Drug Discovery)
- Rapid Publication: manuscripts are peer-reviewed and a first decision is provided to authors approximately 22.8 days after submission; acceptance to publication is undertaken in 4.9 days (median values for papers published in this journal in the second half of 2024).
- Recognition of Reviewers: reviewers who provide timely, thorough peer-review reports receive vouchers entitling them to a discount on the APC of their next publication in any MDPI journal, in appreciation of the work done.
Impact Factor:
3.0 (2023);
5-Year Impact Factor:
4.7 (2023)
Latest Articles
IgG to Galactose-Alpha-1,3-Galactose: Impact of Alpha-Gal IgE Sensitization, Blood Type, and Tick Bites
Antibodies 2025, 14(2), 43; https://doi.org/10.3390/antib14020043 - 16 May 2025
Abstract
Background: Antibodies to galactose-alpha-1,3-galactose (alpha-gal), particularly the IgM and IgG isotypes, are abundant in human sera. These antibodies are known to be an important xenotransplantation barrier, but the full implications of these antibodies to health and disease remain incompletely understood. By contrast, IgE
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Background: Antibodies to galactose-alpha-1,3-galactose (alpha-gal), particularly the IgM and IgG isotypes, are abundant in human sera. These antibodies are known to be an important xenotransplantation barrier, but the full implications of these antibodies to health and disease remain incompletely understood. By contrast, IgE to alpha-gal is uncommon in the population but has been associated with tick bites and causally linked with mammalian meat allergy, often now known as alpha-gal syndrome (AGS). To date, there have been few population-based studies that have investigated alpha-gal IgG levels in relation to demographic factors, diet, tick bites, and mammalian meat allergy. Methods: Adults, predominantly healthcare workers, were recruited for a COVID-19 vaccine study. At least one serum sample was collected, and subjects completed questionnaires to provide demographic, diet, and tick exposure data. Alpha-gal IgG, IgE, and total IgG were measured using the ImmunoCAP platform, and blood group was assessed via reverse typing using stored serum. We also assessed alpha-gal IgG levels among subjects with AGS, recruited from an allergy clinic. Results: The median age of the 267 subjects in the vaccine cohort was 42 years, and median alpha-gal IgG levels were 3.0 μg/mL. Alpha-gal IgG levels were higher among the 43 (16.1%) subjects who had alpha-gal IgE sensitization (≥0.1 IU/mL) and among subjects lacking the B blood group antigen (blood groups A and O). Alpha-gal IgG levels did not differ between the subjects who had asymptomatic alpha-gal IgE sensitization and those who had meat allergy. However, both groups had higher alpha-gal IgG levels than subjects who lacked alpha-gal IgE sensitization. Subjects who reported prior tick or chigger bites had higher alpha-gal IgG levels than those without a bite history, regardless of alpha-gal IgE sensitization status. Conclusions: In a population-based cohort, alpha-gal IgG antibodies were found to be prevalent, and levels were increased in subjects with blood groups A and O, subjects who were alpha-gal IgE sensitized, and those who reported a history of tick bites.
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(This article belongs to the Section Humoral Immunity)
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Open AccessArticle
Neutralization of the Pandemic Influenza A/H1N1 Virus with Lama glama Humanized Nanobodies (VHH)
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Zeila Yazmín Páez-Hernández, Jose Luis Stephano-Hornedo, Jose Alberto Bolaños-Prats, Iván Córdova-Guerrero, Mariana Macías-Alonso, Joaquín G. Marrero, Angel Pulido Capiz and Victor García González
Antibodies 2025, 14(2), 42; https://doi.org/10.3390/antib14020042 - 16 May 2025
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Background/Objetives: Nanobodies (VHH) have become an excellent tool for diagnosis, therapy, and research since VHH shows a high capability of recognizing and neutralizing antigens. VHHs are highly soluble and stable at high temperatures, and in the presence of chaotropic agents, they offer significant
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Background/Objetives: Nanobodies (VHH) have become an excellent tool for diagnosis, therapy, and research since VHH shows a high capability of recognizing and neutralizing antigens. VHHs are highly soluble and stable at high temperatures, and in the presence of chaotropic agents, they offer significant advantages over other biological therapeutic agents. This study aimed to identify and humanize VHH fragments with neutralizing potential against the influenza A/H1N1 virus. Methods: A library of VHH antibody fragments was produced by phage display technique against an inactivated influenza A/H1N1 vaccine. Three VHH sequences were selected and humanized. Specifically, the recognition capacity of the antibodies denominated 2-C10 and 2-C10H was confirmed by ELISA and western blot (WB), as well as their microneutralization capacity in a cellular model, suggesting their potential therapeutic use in patients infected with the influenza A/H1N1 virus. Molecular docking assays were used to support the mechanism of viral inhibition. Results: The VHHs 2-C10 and 2-C10H showed specific recognition of influenza A/H1N1 antigens by ELISA and Western Blot and demonstrated neutralizing activity in vitro. The optimal VHH, 2-C10H, showed 75% neutralization capacity at a concentration of 1.56 μg/mL against the A/H1N1 viral strain, potentially through the inactivation of hemagglutinin protein, a phenomenon supported by molecular docking assays. Conclusions: This study presents a strategic approach to identify VHH candidates that may be useful for diagnosing and potentially treating patients already infected by the A/H1N1 virus, as it may reduce the severity of their symptoms.
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Open AccessArticle
Generative Deep Learning Design of Single-Domain Antibodies Against Venezuelan Equine Encephalitis Virus
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Jinny L. Liu, Gabrielle C. Bayacal, Jerome Anthony E. Alvarez, Lisa C. Shriver-Lake, Ellen R. Goldman and Scott N. Dean
Antibodies 2025, 14(2), 41; https://doi.org/10.3390/antib14020041 - 14 May 2025
Abstract
Background/Objectives: Venezuelan equine encephalitis virus (VEEV) represents a significant biothreat with no FDA-approved vaccine currently available, highlighting the need for alternative therapeutic strategies. Single-domain antibodies (sdAbs) present a potential alternative to conventional antibodies, due to their small size and ability to recognize cryptic
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Background/Objectives: Venezuelan equine encephalitis virus (VEEV) represents a significant biothreat with no FDA-approved vaccine currently available, highlighting the need for alternative therapeutic strategies. Single-domain antibodies (sdAbs) present a potential alternative to conventional antibodies, due to their small size and ability to recognize cryptic epitopes. Methods: This research describes the development and preliminary evaluation of VEEV-binding sdAbs generated using a generative artificial intelligence (AI) platform. Using a dataset of known alphavirus-binding sdAbs, the AI model produced sequences with predicted affinity for the E2 glycoprotein of VEEV. These candidate sdAbs were expressed in a bacterial periplasmic system and purified for initial assessment. Results: Enzyme-linked immunosorbent assays (ELISAs) indicated binding activity of the sdAbs to VEEV antigens. In vitro neutralization tests suggested inhibition of VEEV infection in cultured cells for some of the candidates. Conclusions: This study demonstrates how generative AI can expedite antiviral therapeutic development and establishes a framework for quick responses to emerging viral threats when extensive example databases are unavailable. Additional refinement and validation of AI-generated sdAbs could establish effective VEEV therapeutics.
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(This article belongs to the Section Antibody Discovery and Engineering)
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Open AccessArticle
Purification of Human Immunoglobulin G with Bathophenanthroline–Zn2+, –Fe2+, or –Cu2+ Complexes
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Thisara Jayawickrama Withanage, Ron Alcalay, Olga Krichevsky, Ellen Wachtel, Ohad Mazor and Guy Patchornik
Antibodies 2025, 14(2), 40; https://doi.org/10.3390/antib14020040 - 12 May 2025
Abstract
Background/Objectives: Pharmaceutical companies are aware of the ongoing effort to satisfy the increasing global demand for therapeutic-grade monoclonal antibodies (mAbs), an especially difficult challenge for poor and developing countries. We present a simple, economical, single-step purification approach at neutral pH for polyclonal human
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Background/Objectives: Pharmaceutical companies are aware of the ongoing effort to satisfy the increasing global demand for therapeutic-grade monoclonal antibodies (mAbs), an especially difficult challenge for poor and developing countries. We present a simple, economical, single-step purification approach at neutral pH for polyclonal human IgG (hIgG), which does not require any expensive ligands, chromatography columns, polymers, or membranes. Methods/Results: Instead, porous precipitates of commercial, recyclable aromatic [bathophenanthroline:cation] complexes were found to efficiently capture impurity proteins from CHO cells or E. coli lysate while maintaining the majority of the highly concentrated hIgG (5–15 mg/mL) in the supernatant. [(Batho)3:Zn2+] complexes were the most promising, resulting in hIgG with a purity of ≈95%, by SDS-PAGE. This purified hIgG is monomeric (by dynamic light scattering, DLS) and preserves the native secondary structure (by far UV circular dichroism spectroscopy, CD). The process yield is >90% (by densitometry) and is maintained after a 100-fold increase in the reaction volume, which required only proportional increases in reagents. Conclusions: Although Protein A chromatographic columns, the industry gold standard, have a limited binding capacity, are costly, and require familiarity with column maintenance, we are attempting, by our efforts, to help to produce a more efficient, simple, and economical purification platform.
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(This article belongs to the Section Antibody-Based Therapeutics)
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Open AccessArticle
Validation and Optimization of PURE Ribosome Display for Screening Synthetic Nanobody Libraries
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Bingying Liu and Daiwen Yang
Antibodies 2025, 14(2), 39; https://doi.org/10.3390/antib14020039 - 2 May 2025
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Background/Objectives: PURE (Protein synthesis Using Recombinant Elements), an ideal system for ribosome display, has been successfully used for nanobody selection. However, its limitations in nanobody selection, especially for synthetic nanobody libraries, have not been clearly elucidated, thereby restricting its utilization. Methods: The PURE
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Background/Objectives: PURE (Protein synthesis Using Recombinant Elements), an ideal system for ribosome display, has been successfully used for nanobody selection. However, its limitations in nanobody selection, especially for synthetic nanobody libraries, have not been clearly elucidated, thereby restricting its utilization. Methods: The PURE ribosome display selection process was closely monitored using RNA agarose gel electrophoresis to assess the presence of mRNA molecules in each fraction, including the flow-through, washing, and elution fractions. Additionally, a real-time validation method for monitoring each biopanning round was implemented, ensuring the successful enrichment of target protein-specific binders. The selection process was further optimized by introducing a target protein elution step prior to the EDTA-mediated disassembly, as well as by altering the immobilization surfaces. Finally, the efficiency of PURE ribosome display was enhanced by replacing the spacer gene. Results: The efficiency of PURE ribosome display was merely 4% with an unfavourable spacer gene. Using this spacer gene, EGFP- and human fatty acid-binding protein 4-specific nanobodies from a synthetic nanobody library were we successfully identified through optimizing the selection process. Choosing a spacer gene less prone to secondary structure formation increased significantly its efficiency in displaying synthetic nanobody libraries. Conclusions: Implementing a target protein elution step prior to EDTA-mediated disassembly and modifying the immobilization surfaces effectively increase selection efficiency. For PURE ribosome display, efficiency was further improved using a suitable spacer gene, enabling the display of large libraries.
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Open AccessArticle
A Targeted Integration-Based CHO Cell Platform for Simultaneous Antibody Display and Secretion
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Jessica P. Z. Ng, Mariati Mariati, Jiawu Bi, Matthew Wook Chang and Yuansheng Yang
Antibodies 2025, 14(2), 38; https://doi.org/10.3390/antib14020038 - 28 Apr 2025
Abstract
Objective: We developed a targeted integration-based CHO cell platform for simultaneous antibody display and secretion, enabling a streamlined transition from antibody library screening to production without requiring the re-cloning of antibody genes. Methods: The platform consists of a CHO master cell line with
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Objective: We developed a targeted integration-based CHO cell platform for simultaneous antibody display and secretion, enabling a streamlined transition from antibody library screening to production without requiring the re-cloning of antibody genes. Methods: The platform consists of a CHO master cell line with a single-copy landing pad, a helper vector expressing FLPe recombinase, and bi-functional targeting vectors. Recombinase-mediated cassette exchange was utilized to integrate targeting vectors into the landing pad. Bi-functional vectors were designed by incorporating a minimal furin cleavage sequence (mFCS), RRKR, and various 2A peptides between the heavy chain (HC) and a membrane anchor. Results: Incomplete cleavage at the mFCS and 2A sites facilitated the expression of both membrane-bound and secreted antibodies, while mutations in the 2A peptide produced a range of display-to-secretion ratios. However, a fraction of secreted antibodies retained 2A residues attached to the HC polypeptides. Further analysis demonstrated that modifying the first five amino acids of the 2A peptide significantly influenced furin cleavage efficiency, resulting in different display-to-secretion ratios for targeting vectors containing mFCS-2A variant combinations. To overcome this, we designed nine-amino-acid FCS variants that, when placed between the HC and membrane anchor, provided a range of display-to-secretion ratios and eliminated the issue of attached 2A residues in the secreted antibodies. Vectors with lower display levels proved more effective at distinguishing cells expressing high-affinity antibodies with closely matched binding affinities. The platform also demonstrated high sensitivity in isolating high-affinity antibody-expressing cells and supported robust antibody production. Conclusion: This targeted integration-based CHO platform enables efficient, in-format screening and production of antibodies with tunable display-to-secretion profiles. It provides a powerful and scalable tool for accelerating the development of functional, manufacturable therapeutic antibodies.
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(This article belongs to the Special Issue Emerging Antibody Engineering Strategies and Applications for Immunotherapy of Cancer)
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Open AccessReview
The Role of Monoclonal Antibodies as Therapeutics in HPV-Related Head and Neck Cancers: An Updated Review
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Michael Zalin, Shaan Patel, Carter Coggins and Vikrant Rai
Antibodies 2025, 14(2), 37; https://doi.org/10.3390/antib14020037 - 24 Apr 2025
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Background/Objectives: The increasing prevalence of human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) has necessitated a revaluation of therapeutic strategies. HPV-driven OPSCC differs from HPV-negative OPSCC due to its distinct molecular signatures, increased radiosensitivity, and better prognoses. However, despite these differences, treatment strategies
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Background/Objectives: The increasing prevalence of human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) has necessitated a revaluation of therapeutic strategies. HPV-driven OPSCC differs from HPV-negative OPSCC due to its distinct molecular signatures, increased radiosensitivity, and better prognoses. However, despite these differences, treatment strategies have remained largely uniform, resulting in minimal reductions in morbidity and exposing HPV-positive patients to unnecessary toxicity. Monoclonal antibodies (mAbs) have become a promising therapeutic option due to their ability to target treatment with fewer systemic side effects. Immune checkpoint inhibitors (ICIs) such as pembrolizumab have shown efficacy in enhancing the immune response against tumors, while EGFR inhibitors like cetuximab offer an alternative modality. Current clinical trials aim to refine dosing regimens and identify combination strategies that may enhance therapeutic outcomes. Results: Despite promising evidence, several challenges hinder the widespread adoption of mAbs as a standard treatment for HPV-positive OPSCC in clinical practice. This review examines the current role of mAbs in HPV-positive OPSCC treatment, highlighting their limitations and future research directions. Conclusions: Further studies are needed to optimize patient selection, establish standardized treatment protocols, and investigate the long-term benefits of mAb-based therapies in this patient population.
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Open AccessArticle
Potentiating Antibody-Dependent Cellular Cytotoxicity in Triple-Negative Breast Cancer via the Humanized Anti-CD147 Antibody
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Kanyarat Thongheang, Thanathat Pamonsupornwichit, Kanokporn Sornsuwan, On-anong Juntit, Tawan Chokepaichitkool, Weeraya Thongkum, Umpa Yasamut and Chatchai Tayapiwatana
Antibodies 2025, 14(2), 36; https://doi.org/10.3390/antib14020036 - 11 Apr 2025
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Background: Triple-negative breast cancer (TNBC) is an aggressive subtype with high metastatic potential, poor prognosis, and the absence of estrogen receptors, progesterone receptors, and human epidermal growth factor receptor 2 (HER2). The lack of these receptors limits the standard treatments, such as hormone
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Background: Triple-negative breast cancer (TNBC) is an aggressive subtype with high metastatic potential, poor prognosis, and the absence of estrogen receptors, progesterone receptors, and human epidermal growth factor receptor 2 (HER2). The lack of these receptors limits the standard treatments, such as hormone therapies and HER2-targeted antibodies like trastuzumab. These challenges highlight the critical need for novel therapeutic strategies. CD147, a transmembrane glycoprotein overexpressed in TNBC, promotes tumor progression, metastasis, and chemoresistance, making it a promising therapeutic target. This study evaluates the antibody-dependent cellular cytotoxicity (ADCC) of HuM6-1B9, a humanized anti-CD147 antibody, against MDA-MB-231 cells, a TNBC model. Methods: CFSE-labelled MDA-MB-231 cells were co-cultured with PBMCs as effector cells (E:T ratio 80:1) in the presence of HuM6-1B9 and incubated for 4 h. Cells were then collected and stained with PI, and CFSE+/PI+ dead target cells were analyzed by flow cytometry. Results: Co-culturing MDA-MB-231 cells with peripheral blood mononuclear cells (PBMCs) in the presence of HuM6-1B9 demonstrated effective ADCC induction without direct cytotoxicity. HuM6-1B9 induced 54.01% cancer cell death via ADCC, significantly outperforming trastuzumab (26.14%) while sparing PBMCs. Conclusion: These findings support HuM6-1B9 as a prospective TNBC therapeutic and warrant further investigation into its clinical potential.
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Open AccessReview
A Comprehensive Review About the Use of Monoclonal Antibodies in Cancer Therapy
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Angel Justiz-Vaillant, Bijay Raj Pandit, Chandrashekhar Unakal, Sehlule Vuma and Patrick Eberechi Akpaka
Antibodies 2025, 14(2), 35; https://doi.org/10.3390/antib14020035 - 11 Apr 2025
Abstract
Monoclonal antibodies (mAbs) targeting various pathways in cancer therapy play crucial roles in enhancing the immune system’s ability to recognise and eliminate tumour cells. These therapies are designed to either block inhibitory immune checkpoint pathways or to target specific tumour cell markers for
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Monoclonal antibodies (mAbs) targeting various pathways in cancer therapy play crucial roles in enhancing the immune system’s ability to recognise and eliminate tumour cells. These therapies are designed to either block inhibitory immune checkpoint pathways or to target specific tumour cell markers for direct destruction. Additionally, mAbs can modulate the tumour microenvironment, enhance antibody-dependent cellular cytotoxicity, and inhibit angiogenesis, further amplifying their therapeutic impact. Below is a summary of monoclonal antibodies targeting key pathways, along with their indications and mechanisms of action, which are reviewed based on therapeutic mechanisms.
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(This article belongs to the Special Issue Emerging Antibody Engineering Strategies and Applications for Immunotherapy of Cancer)
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Open AccessCase Report
An Exceedingly Rare Case of Mechanobullous Epidermolysis Bullosa Acquisita in a Prepubertal Child: A Review of the Clinical and Laboratory Considerations
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Aleksandra Wiktoria Bratborska, Maciej Spałek, Monika Bowszyc-Dmochowska and Marian Dmochowski
Antibodies 2025, 14(2), 34; https://doi.org/10.3390/antib14020034 - 11 Apr 2025
Abstract
Introduction: Epidermolysis bullosa acquisita (EBA) is a rare autoimmune disease causing subepithelial blistering due to autoantibodies against type VII collagen. While mechanobullous EBA predominantly affects adults, our report presents an exceedingly rare case in an 11-year-old football player. Case Report: The patient reported
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Introduction: Epidermolysis bullosa acquisita (EBA) is a rare autoimmune disease causing subepithelial blistering due to autoantibodies against type VII collagen. While mechanobullous EBA predominantly affects adults, our report presents an exceedingly rare case in an 11-year-old football player. Case Report: The patient reported a one-year history of blistering and scarring on the knees and scrotum. The diagnosis was established with direct immunofluorescence (DIF), mosaic indirect immunofluorescence (IIF) showing IgG antibodies reacting with the dermal side of salt-split primate skin, and multiplex ELISA revealing an elevated level of IgG antibodies against type VII collagen. Treatment with a superpotent topical glucocorticosteroid and activity modifications improved his condition. Review: This case highlights the importance of considering EBA in differential diagnoses of pediatric blistering diseases and suggests that conservative management may be effective in mild cases. We also review clinical and laboratory considerations on the topic of childhood EBA. Conclusions: Further studies are essential to develop evidence-based guidelines for pediatric EBA.
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(This article belongs to the Section Antibody-Based Diagnostics)
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Open AccessArticle
Effect of Acute Lung Injury (ALI) Induced by Lipopolysaccharide (LPS) on the Pulmonary Pharmacokinetics of an Antibody
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Shweta Jogi and Dhaval K. Shah
Antibodies 2025, 14(2), 33; https://doi.org/10.3390/antib14020033 - 6 Apr 2025
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Objective: To investigate the effect of Lipopolysaccharide (LPS)-induced acute lung injury (ALI) on the pulmonary pharmacokinetics (PK) of a systemically administered antibody in mice. Method: The PK of a non-target-binding antibody was evaluated in healthy mice and mice with intratracheal instillation of 5
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Objective: To investigate the effect of Lipopolysaccharide (LPS)-induced acute lung injury (ALI) on the pulmonary pharmacokinetics (PK) of a systemically administered antibody in mice. Method: The PK of a non-target-binding antibody was evaluated in healthy mice and mice with intratracheal instillation of 5 mg/kg LPS. The plasma, bronchoalveolar lavage (BAL), trachea, bronchi, and lung homogenate PK of the antibody were measured following intravenous administration of 5 mg/kg antibody dose. Noncompartmental analysis was performed to determine AUC values. Antibody concentrations in all biological matrices were quantified using qualified ELISA. The effect of ALI on BAL albumin and total protein concentrations was also determined. BAL protein concentrations were corrected for dilution using plasma urea concentrations. Results: Intratracheal instillation of LPS and the resultant ALI led to ~2–4-fold higher concentrations of albumin and proteins in the BAL. LPS-induced ALI also notably altered the pulmonary PK of the antibody. The effect of ALI on the antibody PK was time and tissue dependent. The trachea and bronchi showed ~1.7-fold and ~1.4-fold lower antibody exposure compared with the control group, but the BAL fluid exhibited ~4-fold increase in antibody exposure following LPS treatment. Most noticeable changes in antibody PK occurred 24 h after LPS administration, and the effect was temporary for the bronchi and trachea. However, the changes in lung homogenate and, more notably, in BAL persisted until the end of the experiment. Thus, our investigation suggests that due to the acute nature of ALI-induced pathophysiology and the changing severity of the disease, the dose and timing of antibody administration following ALI may need to be optimized based on the target site of action (e.g., bronchi, trachea, BAL, lung parenchyma, etc.) to maximize the therapeutic effect of the antibody. Conclusions: ALI may significantly affect pulmonary PK of systemically administered antibodies. Changes caused by ALI are time and tissue dependent, and hence, the timing and dose of antibody following ALI may need to be optimized to maximize the therapeutic effect of the antibody at the site of action.
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Open AccessArticle
A Nationwide Seroprevalence Study for Measles in Individuals of Fertile Age in Romania
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Aurora Stanescu, Simona Maria Ruta, Mihaela Leustean, Ionel Iosif, Camelia Sultana, Anca Maria Panaitescu, Florentina Ligia Furtunescu, Costin Cernescu and Adriana Pistol
Antibodies 2025, 14(2), 32; https://doi.org/10.3390/antib14020032 - 2 Apr 2025
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Background/Objectives: Romania remains endemic for measles due to suboptimal vaccine coverage rates. During the last three epidemics, the highest incidence of measles was recorded in children younger than 1 year, who should have been partially protected by maternal antibodies. A nationwide cross-sectional seroprevalence
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Background/Objectives: Romania remains endemic for measles due to suboptimal vaccine coverage rates. During the last three epidemics, the highest incidence of measles was recorded in children younger than 1 year, who should have been partially protected by maternal antibodies. A nationwide cross-sectional seroprevalence study was conducted on persons of fertile age, to evaluate potential immunity gaps in the population. Methods: Between June and October 2020, 959 serum samples were collected from individuals aged 25–44 years (46.5% females) from all the geographic regions in Romania. Measles IgG antibodies were assessed using an enzyme-linked immune assay (DIA.PRO-Diagnostic Bioprobes Srl, Italy). Statistical analysis was performed in IBM SPSS Statistics 27.0, using Fisher’s exact and chi-squared tests to test for associations between seropositivity and demographic factors, with p < 0.05 considered statistically significant. Results: The overall measles seroprevalence was 77%, without gender- or geographic region-related differences. Both the seropositivity rate and the measles antibodies titers increased with age, with the highest difference between the oldest and the youngest age group (p = 0.057), suggesting persistent immunity after natural infection in older individuals or anamnestic responses in vaccinated persons, caused by repeated exposures to the circulating virus. An additional confirmatory pilot study on 444 pregnant women confirmed the low level of measles seroprevalence (68.4%), with a significant upward trend in older ages (75% in those aged >40 years old vs. 65% in those aged 25–29 years, p = 0.018 and mean reactivity of measles antibodies 3.05 ± 1.75 in those aged >40 years vs. 2.28 ± 1.39 in those aged 25–29 years, p = 0.037). Conclusions: This study signals critical immunity gaps in the population that contribute to the accumulation of susceptible individuals and recurrent measles outbreaks. The absence of measles antibodies in women of childbearing age increases the newborn’s susceptibility to infection, with potentially severe complications.
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Open AccessArticle
Impact of Monoclonal Antibody Aggregates on Effector Function Characterization
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Wendy J. Walton, Shousong Jason Zhang, Joseph J. Wilson, Briana N. Harvey, Matthew Clemens and Yingmei Gu
Antibodies 2025, 14(2), 31; https://doi.org/10.3390/antib14020031 - 2 Apr 2025
Abstract
Background/Objectives: Monoclonal antibodies have successfully been used for a variety of indications. Many therapeutic antibodies are IgG1 and elicit effector functions as part of their mechanism of action. It is well known that aggregate levels should be controlled for therapeutic antibodies. Although there
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Background/Objectives: Monoclonal antibodies have successfully been used for a variety of indications. Many therapeutic antibodies are IgG1 and elicit effector functions as part of their mechanism of action. It is well known that aggregate levels should be controlled for therapeutic antibodies. Although there are several reports describing the impact of antibody aggregates on FcγR binding, most of these have been performed with surface plasmon resonance in an avidity-based format. What is less well known is which Fcγ receptor is most impacted by antibody aggregation and how antibody aggregates impact binding to Fcγ receptors in solution-based formats and in cell-based assays. Methods: An effector-competent IgG1 (mAb1) was forcibly degraded and fractionated by size exclusion chromatography to enrich for aggregates. The fractions were examined for FcγR binding by SPR with different formats and in solution. The fractions were also analyzed with cell-based FcγR reporter assays. Results: All Fcγ receptors displayed increased binding to enriched mAb1 aggregates in the avidity-based SPR methods and in solution, with FcγRIIa impacted the most. When examined with an antibody-down SPR format that is not usually susceptible to avidity, FcγRIIa did not show increased binding with mAb1 aggregation. Although activity for mAb1 aggregates increased slightly in an FcγRIIa cell-based reporter assay, it decreased in the FcγRIIIa reporter assay (most likely due to differences in fucosylation from the reference standard). Conclusions: Monoclonal antibody aggregation can impact FcγR binding for avidity-based binding formats. Even at low levels of antibody aggregation, FcγRII binding increases substantially.
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(This article belongs to the Section Antibody-Based Therapeutics)
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Open AccessArticle
T330M Substitution in the Sodium-Dependent Phosphate Transporter NaPi2b Abolishes the Efficacy of Monoclonal Antibodies Against MX35 Epitope
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Leisan F. Bulatova, Vera S. Skripova, Aisylu R. Sagdeeva, Ramilia A. Vlasenkova, Tatiana A. Bugaenko, Rezeda R. Galimova, Alfiya I. Nesterova, Yuliya V. Filina and Ramziya G. Kiyamova
Antibodies 2025, 14(2), 30; https://doi.org/10.3390/antib14020030 - 1 Apr 2025
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Background: Monoclonal antibodies against the sodium-dependent phosphate transporter NaPi2b (SLC34A2) represent a promising approach in the treatment of ovarian and lung cancer. Of particular interest is the potential cancer-specific MX35 epitope of NaPi2b, as it serves as a target for monoclonal
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Background: Monoclonal antibodies against the sodium-dependent phosphate transporter NaPi2b (SLC34A2) represent a promising approach in the treatment of ovarian and lung cancer. Of particular interest is the potential cancer-specific MX35 epitope of NaPi2b, as it serves as a target for monoclonal antibodies studied at various stages of preclinical and clinical trials. However, variations in the NaPi2b protein structure may limit the efficacy of therapeutic antibodies by affecting the accessibility of the MX35 epitope. Methods: An in silico analysis was performed using data from 101,562 tumor samples. Genomic DNA sequencing was conducted on blood samples from patients with ovarian carcinoma, breast cancer, and renal carcinoma to access the frequency of germline mutations in the SLC34A2 gene region encoding the MX35 epitope. To assess the impact of the selected mutation, we generated a model cell line through site-directed mutagenesis carrying the mutant NaPi2b variant. Results: Using in silico analysis, we identified 17 unique variants in the SLC34A2 gene leading to amino acid substitutions within the MX35 epitope of the NaPi2b. Among these, the most prevalent mutation, c.989C>T, resulting in p.T330M substitution, was detected in 5 out of 64 patients through genomic DNA sequencing. Using site-directed mutagenesis, we created the OVCAR-8/NaPi2bp.T330M model cell line. L3 (28/1) monoclonal antibodies specific to the MX35 epitope failed to recognize the mutant NaPi2bp.T330M variant compared to the wild-type of the NaPi2b in both Western blot and confocal microscopy experiments. Conclusions: The obtained data may serve as a basis for predicting the efficacy of monoclonal antibody-based targeted therapy binding to the MX35 epitope of NaPi2b in the treatment of oncological diseases.
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Open AccessArticle
A Novel Method for Preparing Uniform Micro-Sized Dry Powder Formulations, Including Aggregation-Controlled VHH
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Tatsuru Moritani, Hidekazu Masaki, Ryo Yonehara, Takeru Suzuki, Hidenao Arai, Masayuki Tsuchiya and Naoto Nemoto
Antibodies 2025, 14(2), 29; https://doi.org/10.3390/antib14020029 - 31 Mar 2025
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Background: The preparation of antibodies in powder form without changing their physicochemical properties may enable their use in new drug delivery system therapies or non-refrigerated storage. The variable domain of heavy-chain antibodies (VHHs) is more suited for this purpose than that of conventional
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Background: The preparation of antibodies in powder form without changing their physicochemical properties may enable their use in new drug delivery system therapies or non-refrigerated storage. The variable domain of heavy-chain antibodies (VHHs) is more suited for this purpose than that of conventional antibodies because of VHHs’ high thermal stability and ability to refold. Methods: In this report, the fine droplet drying (FDD) process was selected as the powderization technique because of its favorable features, such as mild drying conditions and the generation of uniform particle sizes. The aggregation, binding, particle, and in vitro inhalation properties of the prepared VHH powders (VHHps) were evaluated. Results: The amount of aggregated VHHs present in the VHHps depended on the flow temperature during the FDD process, with higher temperatures yielding a higher aggregation ratio. In contrast, no significant difference in binding activity was observed between each VHHp preparation and the native VHHs. However, this process degraded VHHs or inactivated their function, and ultimately, only about 30% of the original VHHs were functional, whereas the remaining VHHs that were not degraded showed little loss of functionality, even after storage at room temperature for more than two years. Analysis of the VHHp samples revealed that the particles were uniformly spherical with a single-micron size. The VHHps showed fine inhalation properties in the inhalation property test. Conclusions: These findings suggest that the FDD process affords various VHH powder formulations, including pharmaceutical formulations.
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Open AccessArticle
Discovery of Antibodies Against Endemic Coronaviruses with NGS-Based Human Fab Phage Display Platform
by
Oscar Chi-Chien Pan, Sean Miller, Ruchin Patel, Shreya Mukhopadhyay, Giancarlo Sarullo, Gwenny Go, Jennifer Galli, Jamie Hessels, Barbara Schlingmann-Molina, Emmanuel Ndashimye, Zhiyun Wen, Christopher Warren, Eberhard Durr, Lan Zhang, Kalpit A. Vora, Arthur Fridman and Zhifeng Chen
Antibodies 2025, 14(2), 28; https://doi.org/10.3390/antib14020028 - 27 Mar 2025
Cited by 1
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Background: There is an unmet medical need to develop a vaccine targeting endemic coronaviruses. Antigen-specific monoclonal antibodies (mAbs) are crucial for many assays to support vaccine development. Objective: In this study, we used the HuCal Fab phage display library with a diversity of
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Background: There is an unmet medical need to develop a vaccine targeting endemic coronaviruses. Antigen-specific monoclonal antibodies (mAbs) are crucial for many assays to support vaccine development. Objective: In this study, we used the HuCal Fab phage display library with a diversity of 4.5 × 1010 to identify antibodies specific to the spike proteins of the four endemic coronaviruses: OC43, NL63, 229E, and HKU1. Methods: As proof of concept, we established a newly designed platform using a long-read NGS workflow for antibody discovery and compared the results against the traditional workflow using Sanger sequencing consisting of lengthy and laborious benchwork. Results: The long-read NGS workflow identified most of the antibodies seen from the Sanger sequencing workflow, and many more additional antigen-specific antibodies against the endemic coronaviruses. Overall efficiency improved up to three times, comparing the traditional workflow with the NGS workflow. Of the 113 NGS-derived mAbs isolated to bind the four endemic coronavirus spike proteins, 107/113 (94.7%) had potent ELISA binding affinities (EC50 < 150 ng/mL, or <1 nM), and 61/113 (54%) had extremely potent ELISA binding affinities (EC50 of <15 ng/mL, or <0.1 nM). Conclusions: We successfully developed and incorporated the long-read NGS workflow to generate target-specific antibodies with many antibodies at sub-nanomolar affinities that are likely missed by a traditional workflow. We identified strong neutralizing antibodies, proving that our endemic spike proteins are capable of generating antibodies that could offer protection against the endemic HCoVs.
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Open AccessArticle
Low Serological Agreement of Hepatitis E in Immunocompromised Cancer Patients: A Comparative Study of Three Anti-HEV Assays
by
Isabel-Elena Haller, Mark Reinwald, Janine Kah, Franz A. M. Eggert, Sandra Schwarzlose-Schwarck, Kristoph Jahnke, Stefan Lüth and Werner Dammermann
Antibodies 2025, 14(2), 27; https://doi.org/10.3390/antib14020027 - 24 Mar 2025
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Background/Objectives: Hepatitis E virus (HEV) is one of the leading causes of acute hepatitis, with immunosuppressed individuals, such as oncology patients, being particularly vulnerable to chronic infections that may progress to liver disease or fatal outcomes. Assay variability complicates HEV prevalence assessment in
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Background/Objectives: Hepatitis E virus (HEV) is one of the leading causes of acute hepatitis, with immunosuppressed individuals, such as oncology patients, being particularly vulnerable to chronic infections that may progress to liver disease or fatal outcomes. Assay variability complicates HEV prevalence assessment in at-risk groups. This study aimed to compare the reliability and concordance of three HEV antibody assays—Wantai, Euroimmun, and Elecsys®—in immunosuppressed oncology patients. Methods: In this prospective pilot study, serum samples were obtained from oncology patients between September 2020 and October 2021. Samples were collected both at baseline (treatment-naive) and during ongoing treatment. A healthy control group was retrospectively included for comparative analysis. Anti-HEV IgM and IgG antibodies were tested in all samples using enzyme-linked immunosorbent assays (Wantai, Euroimmun) and an electrochemiluminescence immunoassay (Elecsys®). Demographic and clinical data, along with information on HEV risk factors, were extracted from medical records and patient questionnaires. Results: HEV IgM prevalence ranged from 0% (Wantai) to 6% (Elecsys®), while IgG prevalence was 12% (Euroimmun), 38% (Wantai), and 53% (Elecsys®). Concordance was poor, with Cohen’s Kappa values indicating slight to moderate agreement (κ = 0.000–0.553). Patients with hematological malignancies exhibited the highest IgG seroprevalence. Risk factor analysis revealed the highest association between HEV exposure and the consumption of undercooked pork or crop-based agriculture. Conclusions: Significant variability among HEV serological assays highlights the challenges of reliable HEV diagnostics in immunosuppressed oncology patients. Assay selection and improved testing strategies are critical for this high-risk group.
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Open AccessReview
Future Landscape of Anti-Claudin 18.2 Antibodies in Gastric Adenocarcinoma
by
Wendy M. Covert and Jane E. Rogers
Antibodies 2025, 14(1), 26; https://doi.org/10.3390/antib14010026 - 18 Mar 2025
Abstract
Advanced gastric adenocarcinoma (GAC) carries a poor prognosis. Targeted therapy in GAC has traditionally been limited to anti-human epidermal growth factor receptor-2 and anti-vascular endothelial growth factor agents. Recent years have brought immune checkpoint therapy to the GAC treatment landscape. However, continued discovery
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Advanced gastric adenocarcinoma (GAC) carries a poor prognosis. Targeted therapy in GAC has traditionally been limited to anti-human epidermal growth factor receptor-2 and anti-vascular endothelial growth factor agents. Recent years have brought immune checkpoint therapy to the GAC treatment landscape. However, continued discovery of targeted therapy in GAC is needed. Claudins, transmembrane proteins located in tight junctions of epithelial and endothelial cells, help regulate cellular polarity. Claudin dysregulation has been linked to cancers and other diseases. Claudin 18.2 specifically has become a new novel and exciting biomarker for GAC. Many agents are in the investigative pipeline, including monoclonal antibodies, antibody-drug conjugates, bispecific antibodies, and chimeric T-cell therapy. Recently, zolbetuximab, an anti-claudin 18.2 monoclonal antibody, was the first of these agents to get FDA approval. Here, we review zolbetuximab’s place in therapy along with other agents being explored.
Full article
Open AccessReview
Is There a Diagnostic and Prognostic Role for Anti-Nephrin Autoantibodies in Diabetic Nephropathy?
by
Han Sean Lee, Henry H. L. Wu, Arvind Ponnusamy, Helen Alderson and Rajkumar Chinnadurai
Antibodies 2025, 14(1), 25; https://doi.org/10.3390/antib14010025 - 12 Mar 2025
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Diabetic nephropathy (DN) is one of the key causes of end-stage kidney disease worldwide, especially in developed countries. The classic pathogenic development of DN is characterized by microalbuminuria which would progress to nephrotic-range proteinuria and loss of kidney function. The degree of albuminuria
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Diabetic nephropathy (DN) is one of the key causes of end-stage kidney disease worldwide, especially in developed countries. The classic pathogenic development of DN is characterized by microalbuminuria which would progress to nephrotic-range proteinuria and loss of kidney function. The degree of albuminuria is considered an independent risk factor for all-cause mortality in patients with DN. It is now well established that albuminuria stems from disruptions in podocyte structure and function. Podocytes play a major role in the glomerular filtration barrier. The nephrin protein has been identified as a core component of the slit diaphragm in podocytes, and as such, the downregulation of nephrin expression has been described well in various proteinuric glomerulopathies, including DN. Previous studies have shown that the presence of urinary nephrin potentially signifies an early marker of podocyte injury in DN. More recently, there have been increasing bodies of evidence which suggest that circulating autoantibodies targeting nephrin contributes to the pathogenesis of podocytopathies. However, the functional significance of these circulating autoantibodies in patients with DN is not well understood. In this review, we aim to evaluate the significance of nephrin dysregulation in the pathogenesis of DN based on the current available literature and provide an overview on the application of circulating anti-nephrin autoantibodies in relation to its diagnostic as well as prognostic role in podocytopathies, including DN.
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Open AccessReview
Anti-ADAMTS13 Autoantibodies in Immune-Mediated Thrombotic Thrombocytopenic Purpura
by
Michael R. Snyder and Robert W. Maitta
Antibodies 2025, 14(1), 24; https://doi.org/10.3390/antib14010024 - 10 Mar 2025
Cited by 1
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Autoantibodies to ADAMTS13 are at the center of pathology of the immune-mediated thrombotic thrombocytopenic purpura. These autoantibodies can be either inhibitory (enzymatic function) or non-inhibitory, resulting in protein depletion. Under normal physiologic conditions, antibodies are generated in response to foreign antigens, which can
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Autoantibodies to ADAMTS13 are at the center of pathology of the immune-mediated thrombotic thrombocytopenic purpura. These autoantibodies can be either inhibitory (enzymatic function) or non-inhibitory, resulting in protein depletion. Under normal physiologic conditions, antibodies are generated in response to foreign antigens, which can include infectious agents; however, these antibodies may at times cross-react with self-epitopes. This is one of the possible mechanisms mediating formation of anti-ADAMTS13 autoantibodies. The process known as “antigenic mimicry” may be responsible for the development of these autoantibodies that recognize and bind cryptic epitopes in ADAMTS13, disrupting its enzymatic function over ultra large von Willebrand factor multimers, forming the seeds for platelet activation and microthrombi formation. In particular, specific amino acid sequences in ADAMTS13 may lead to conformational structures recognized by autoantibodies. Generation of these antibodies may occur more frequently among patients with a genetic predisposition. Conformational changes in ADAMTS13 between open and closed states can also constitute the critical change driving either interactions with autoantibodies or their generation. Nowadays, there is a growing understanding of the role that autoantibodies play in ADAMTS13 pathology. This knowledge, especially of functional qualitative differences among antibodies and the ADAMTS13 sequence specificity of such antibodies, may make possible the development of targeted therapeutic agents to treat the disease. This review aims to present what is known of autoantibodies against ADAMTS13 and how their structure and function result in disease.
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