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Molecules (Basel, Switzerland) Jan 2024Antibody arrays play a pivotal role in the detection and quantification of biomolecules, with their effectiveness largely dependent on efficient protein immobilization....
Antibody arrays play a pivotal role in the detection and quantification of biomolecules, with their effectiveness largely dependent on efficient protein immobilization. Traditional methods often use heterobifunctional cross-linking reagents for attaching functional residues in proteins to corresponding chemical groups on the substrate surface. However, this method does not control the antibody's anchoring point and orientation, potentially leading to reduced binding efficiency and overall performance. Another method using anti-antibodies as intermediate molecules to control the orientation can be used but it demonstrates lower efficiency. Here, we demonstrate a site-specific protein immobilization strategy utilizing AEP1 (asparaginyl endopeptidase) for building a nanobody array. Moreover, we used a nanobody-targeting enhanced green fluorescent protein (eGFP) as the model system to validate the protein immobilization method for building a nanobody array. Finally, by rapidly enriching eGFP, this method further highlights its potential for rapid diagnostic applications. This approach, characterized by its simplicity, high efficiency, and specificity, offers an advancement in the development of surface-modified protein arrays. It promises to enhance the sensitivity and accuracy of biomolecule detection, paving the way for broader applications in various research and diagnostic fields.
Topics: Antibodies; Cross-Linking Reagents
PubMed: 38257279
DOI: 10.3390/molecules29020366 -
Human Vaccines & Immunotherapeutics Nov 2021Antibodies can provide antiviral protection through neutralization and recruitment of innate effector functions through the Fc domain. While neutralization has long been... (Review)
Review
Antibodies can provide antiviral protection through neutralization and recruitment of innate effector functions through the Fc domain. While neutralization has long been appreciated for its role in antibody-mediated protection, a growing body of work indicates that the antibody Fc domain also significantly contributes to antiviral protection. Recruitment of innate immune cells such as natural killer cells, neutrophils, monocytes, macrophages, dendritic cells and the complement system by antibodies can lead to direct restriction of viral infection as well as promoting long-term antiviral immunity. Monoclonal antibody therapeutics against viruses are increasingly incorporating Fc-enhancing features to take advantage of the Fc domain, uncovering a surprising breadth of mechanisms through which antibodies can control viral infection. Here, we review the recent advances in our understanding of antibody-mediated innate immune effector functions in protection from viral infection and review the current approaches and challenges to effectively leverage innate immune cells via antibodies.
Topics: Antibodies, Monoclonal; Antibodies, Neutralizing; Antibodies, Viral; Antiviral Agents; Immunoglobulin Fc Fragments
PubMed: 34613865
DOI: 10.1080/21645515.2021.1976580 -
Viral Immunology Sep 2017Intravenous immunoglobulin (IVIG) is used to treat or prevent severe viral infection, especially cytomegalovirus (CMV) infections. IVIG was characterized to understand...
Intravenous immunoglobulin (IVIG) is used to treat or prevent severe viral infection, especially cytomegalovirus (CMV) infections. IVIG was characterized to understand its interaction with CMV-infected cells. IVIG retarded CMV spread and reduced virus yields depending on the neutralizing (NT) antibody titer. Immediate early protein synthesis was reduced by IVIG in 3 to 15 h, and IVIG specifically reduced the ratio of 66/68k protein synthesis among immediate early proteins in an NT antibody-dependent manner between 4 and 8 h after infection, indicating that antigenic modulation of CMV-infected cells by IVIG reduced viral protein synthesis and virus production. The half-life of antibody bound to CMV-infected cells was 3.8 h. NT antibody titers to varicella-zoster virus (VZV) and CMV in IVIG were dose dependently absorbed by cells infected with VZV and CMV, respectively, but the antibody titers to CMV and VZV, respectively, were not affected. NT antibody in 0.3 mL of IVIG (15 mg) was specifically absorbed by 10 CMV-infected cells and 10 VZV-infected cells, suggesting that the NT antibody in IVIG might be inactivated by one-tenth of a similar volume of CMV-infected or VZV-infected cells. Various antiviral activities of IVIG may contribute to control and alleviation of CMV infection.
Topics: Antibodies, Neutralizing; Antibodies, Viral; Antigenic Modulation; Antiviral Agents; Cells, Cultured; Cytomegalovirus; Cytomegalovirus Infections; Humans; Immediate-Early Proteins; Immunoglobulin G; Immunoglobulins, Intravenous; Neutralization Tests; Virus Release
PubMed: 28598267
DOI: 10.1089/vim.2016.0151 -
Archives of Pathology & Laboratory... Jun 2000Human anti-mouse antibodies (HAMA) are human immunoglobulins with specificity for mouse immunoglobulins. This topic currently is of interest because of the increased use... (Review)
Review
Human anti-mouse antibodies (HAMA) are human immunoglobulins with specificity for mouse immunoglobulins. This topic currently is of interest because of the increased use of monoclonal mouse antibodies as diagnostic reagents both for in vitro laboratory measurements and for in vivo imaging studies. Monoclonal mouse antibodies also are being used therapeutically. This short article reviews the production of HAMA in patients receiving monoclonal antibodies and illustrates the potential ways that HAMA can interfere with immunoassay measurements. Methods for measuring and neutralizing HAMA also are discussed.
Topics: Animals; Antibodies; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal; Antibody Formation; Antibody Specificity; Humans; Immunoassay; Mice; Reproducibility of Results
PubMed: 10835540
DOI: 10.5858/2000-124-0921-HAMA -
BMC Pregnancy and Childbirth Aug 2022Human milk contains antibodies against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) which may serve as a protective factor through passive immunization...
OBJECTIVE
Human milk contains antibodies against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) which may serve as a protective factor through passive immunization in infants. The objective of this study was to measure the levels of anti-SARS-CoV-2 IgG and IgA in human milk and serum after a SARS-CoV-2 infection.
DESIGN
Breast milk and serum samples from 72 lactating mothers with confirmed SARS-CoV-2 asymptomatic or symptomatic infection were collected 1-229 days after the onset of clinical symptoms related to COVID-19. Seventeen mothers with no history of COVID-19 served as a control group. Enzyme-Linked ImmunoSorbent Assay was performed to analyze antibodies against SARS-CoV-2.
RESULTS
SARS-CoV-2-IgA human milk antibodies were detected in mothers and their concentrations were consistently higher than SARS-CoV-2-IgG antibodies. The serum and breastmilk samples of women with COVID-19 was characterized by a higher concentration of anti-RBD IgA and IgG than the serum from the control group without COVID-19. No statistically significant difference was observed between the antibody levels in the serum samples obtained from symptomatic and asymptomatic women exposed to SARS-CoV-2 and between the antibody level and the time from a positive SARS-CoV-2 test result over the period studied.
CONCLUSION
Our results confirm the presence of SARS-CoV-2 IgA and IgG antibodies in the breastmilk of COVID-19 recovered women and the possibility of these antibodies in providing specific immunologic benefits to breastfeeding infants such as protection against the virus transmission and severity of the acquired COVID-19 disease.
Topics: Antibodies, Viral; COVID-19; Female; Humans; Immunoglobulin A; Immunoglobulin G; Lactation; Milk, Human; SARS-CoV-2
PubMed: 35953773
DOI: 10.1186/s12884-022-04945-z -
Expert Review of Clinical Immunology Dec 2017Novel immune globulin (IG) products (RI-002, RI-001) have been designed to provide protection against respiratory syncytial virus (RSV) mediated respiratory illness... (Review)
Review
RI-002, an intravenous immunoglobulin containing high titer neutralizing antibody to RSV and other respiratory viruses for use in primary immunodeficiency disease and other immune compromised populations.
Novel immune globulin (IG) products (RI-002, RI-001) have been designed to provide protection against respiratory syncytial virus (RSV) mediated respiratory illness while at the same time meeting the manufacturing requirements established by FDA for antibody supplementation in immunocompromised subjects. Areas covered: This review covers the manufacture and development of both RI-001 and RI-002, including the selection of plasma donors for IG preparation with high-titers of anti-RSV antibody, in vitro, and preclinical data in the cotton rat model S. hispidus, and clinical trials including Phase II and compassionate use studies of RI-001 and a multi-center, pivotal Phase III study of RI-002 in PIDD patients. Expert commentary: The data demonstrate that RI-002 is efficacious in the prevention and treatment of RSV in preclinical normal and immune suppressed animal models and is safe and efficacious in the treatment of patients with various forms of primary immunodeficiency disease (PIDD). This product offers potential advantages over other available IG's for prophylaxis in immunocompromised patients requiring polyclonal immunoglobulin supplementation because of its unique antibody composition. In addition to its enhanced neutralizing anti-RSV activity and its polyclonal IG composition, there is preclinical data to support the use of RI-002 for humoral protection against other respiratory pathogens.
Topics: Animals; Antibodies, Neutralizing; Antibodies, Viral; Clinical Trials as Topic; Disease Models, Animal; Expert Testimony; Humans; Immunocompromised Host; Immunoglobulins, Intravenous; Immunologic Deficiency Syndromes; Immunotherapy; Rats; Respiratory Syncytial Virus Infections; Respiratory Syncytial Virus, Human; Sigmodontinae
PubMed: 29035131
DOI: 10.1080/1744666X.2017.1389647 -
Clinical and Experimental Immunology Jul 2022Increasing evidence has linked the humoral immune response with the development of various cancers. Therefore, there is growing interest in investigating the predictive...
Increasing evidence has linked the humoral immune response with the development of various cancers. Therefore, there is growing interest in investigating the predictive value of antibodies to assess overall and tissue site-specific cancer risk. Given the large amount of antibody types and the broad scope of the search (i.e. cancer risk), the primary aim of this systematic review was to present an overview of the most researched antibodies (i.e. immunoglobulin (Ig) isotypes (IgG, IgM, IgA, and IgE), tumour and self-antigen-reactive antibodies, infection-related antibodies) in relation to overall and site-specific cancer risk. We identified various antibody types that have been associated with the risk of cancer. While no significant associations were found for IgM serum levels, studies found an inconsistent association among IgE, IgA, and IgG serum levels in relation to cancer risk. When evaluating antibodies against infectious agents, most studies reported a positive link with specific cancers known to be associated with the specific agent recognized by serum antibodies (i.e. helicobacter pylori and gastric cancer, hepatitis B virus and hepatocellular carcinoma, and human papillomavirus and cervical cancer). Several reports identified autoantibodies, as single biomarkers (e.g. anti-p53, anti-MUC1, and anti-CA125) but especially in panels of multiple autoantibodies, to have potential as diagnostic biomarkers for specific cancer types. Overall, there is emerging evidence associating certain antibodies to cancer risk, especially immunoglobulin isotypes, tumour-associated antigen-specific, and self-reactive antibodies. Further experimental studies are necessary to assess the efficacy of specific antibodies as markers for the early diagnosis of cancer.
Topics: Autoantibodies; Biomarkers; Enzyme-Linked Immunosorbent Assay; Humans; Immunoglobulin A; Immunoglobulin E; Immunoglobulin G; Immunoglobulin M; Neoplasms
PubMed: 35380164
DOI: 10.1093/cei/uxac030 -
MAbs 2023The binding properties of bispecific antibodies (bsAb) are crucial for their function, especially when two antigens are targeted on the same cell surface. Dynamic...
The binding properties of bispecific antibodies (bsAb) are crucial for their function, especially when two antigens are targeted on the same cell surface. Dynamic interactions between each of the antibody's arms and its cognate target cause the formation and decay of a biologically functional ternary complex. How association and dissociation processes work cooperatively, and how they influence the avidity of the ternary complex, is still poorly understood. Here, we present a biosensor assay for the simultaneous measurement of the binding kinetics of the therapeutic bsAb emicizumab (Hemlibra®) and its two targets, the blood coagulation factors IX and X (FIX, FX). We describe an automated workflow to characterize binary and ternary-binding modes, utilizing a Y-shaped DNA nanostructure to immobilize the antigens on a sensor and to emulate conditions on a cell or platelet surface by presenting the antigens with optimal accessibility for the bsAb flown over the sensor as analyte. We find that emicizumab binds FX much stronger than FIX (K = 0.05 µM vs. 5 µM, t = 20 s vs. 1 s) with profound consequences on the avidity of the ternary complex, which is dominated by FX's binding properties and a hand-off mechanism from FX to FIX. Moreover, formation and decay of the ternary complex depend on the bsAb concentration during the association phase. Emicizumab's in-vivo mode of action and the catalytic activation of FX can be rationalized from the analyzed binding kinetics. The assay and workflow are well suited for the screening of bispecific binders in drug discovery and provide valuable new kinetic information. bsAb: bispecific antibody; FVIII/FIX/FX: coagulation factors VIII/IX/X; SPR: surface plasmon resonance; k: association rate constant; k: dissociation rate constant; K: equilibrium dissociation constant; t: dissociation half-life.
Topics: Antibodies, Bispecific; Kinetics; Antibodies, Monoclonal, Humanized; Half-Life
PubMed: 36453702
DOI: 10.1080/19420862.2022.2149053 -
The Journal of Infectious Diseases Sep 2022Passive antibody immunotherapeutics directed against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are promising countermeasures for protection and...
Increased Antibody Avidity and Cross-Neutralization of Severe Acute Respiratory Syndrome Coronavirus 2 Variants by Hyperimmunized Transchromosomic Bovine-Derived Human Immunoglobulins for Treatment of Coronavirus Disease 2019.
Passive antibody immunotherapeutics directed against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are promising countermeasures for protection and treatment of coronavirus disease 2019 (COVID-19). SARS-CoV-2 variants of concern (VOCs) and variants of interest (VOIs) can impact the clinical efficacy of immunotherapeutics. A fully human polyclonal antibody immunotherapeutic purified from plasma of transchromosomic (Tc) bovines hyperimmunized with SARS-CoV-2 WA-1 spike (SAB-185) is being assessed for efficacy in a phase 2/3 clinical trial when different circulating SARS-CoV-2 variants predominated. We evaluated antibody binding, avidity maturation, and SARS-CoV-2 VOCs/VOIs virus-neutralizing capacity of convalescent plasma compared with different lots of SAB-185 and individual Tc bovine sera sequentially obtained after each vaccination against Alpha, Epsilon, Iota, Gamma, Beta, Kappa, and Delta variants. In contrast to convalescent plasma, sera and SAB-185 derived from hyperimmunized Tc bovines demonstrated higher antibody avidity and more potent cross-neutralizing activity of VOCs/VOIs. Thus, SAB-185 is a potential promising therapeutic candidate for the treatment of patients infected with SARS-CoV-2 variants.
Topics: Animals; Antibodies, Neutralizing; Antibodies, Viral; Antibody Affinity; COVID-19; Cattle; Humans; Immunization, Passive; Immunoglobulin G; Neutralization Tests; SARS-CoV-2; Spike Glycoprotein, Coronavirus; COVID-19 Serotherapy
PubMed: 35106573
DOI: 10.1093/infdis/jiac031 -
Current Opinion in Virology Apr 2015The development of accurate tools for predicting B-cell epitopes is important but difficult. Traditional methods have examined which regions in an antigen are likely... (Review)
Review
The development of accurate tools for predicting B-cell epitopes is important but difficult. Traditional methods have examined which regions in an antigen are likely binding sites of an antibody. However, it is becoming increasingly clear that most antigen surface residues will be able to bind one or more of the myriad of possible antibodies. In recent years, new approaches have emerged for predicting an epitope for a specific antibody, utilizing information encoded in antibody sequence or structure. Applying such antibody-specific predictions to groups of antibodies in combination with easily obtainable experimental data improves the performance of epitope predictions. We expect that further advances of such tools will be possible with the integration of immunoglobulin repertoire sequencing data.
Topics: Animals; Antibodies; Computational Biology; Epitopes, B-Lymphocyte; Humans; Immunoglobulins; Sequence Analysis, DNA
PubMed: 25837466
DOI: 10.1016/j.coviro.2015.03.012