-
Immunohematology Jun 2021The Kidd-null phenotype, Jk(a-b-), is rare, and a patient with this phenotype may develop anti-Jk3, a red blood cell (RBC) antibody reactive with a domain common to both... (Review)
Review
The Kidd-null phenotype, Jk(a-b-), is rare, and a patient with this phenotype may develop anti-Jk3, a red blood cell (RBC) antibody reactive with a domain common to both Jk and Jk. Like other antibodies to high-prevalence antigens, the presence of this antibody poses challenges in the immunohematologic evaluation of these patients. Thoughtful laboratory testing is necessary to resolve the antibody specificity and to reveal other underlying antibodies. Moreover, the rarity of the Kidd-null phenotype makes finding blood donors difficult for those who need transfusion and have developed anti-Jk3. This review describes methods used in identifying anti-Jk3 in four pregnant patients. Blood bank records were retrospectively reviewed to illustrate the common approach in anti-Jk3 identification. In all cases, pertinent blood bank history was gathered, and extended RBC phenotyping was performed, followed by adsorption studies and testing of selected RBCs. Underlying antibodies were found in two of the cases. This review also reiterates some common challenges encountered with Kidd antibody analysis and highlights the importance of patient ethnic ancestry and obtaining accurate patient transfusion history. The Kidd-null phenotype, Jk(a–b–), is rare, and a patient with this phenotype may develop anti-Jk3, a red blood cell (RBC) antibody reactive with a domain common to both Jk and Jk. Like other antibodies to high-prevalence antigens, the presence of this antibody poses challenges in the immunohematologic evaluation of these patients. Thoughtful laboratory testing is necessary to resolve the antibody specificity and to reveal other underlying antibodies. Moreover, the rarity of the Kidd-null phenotype makes finding blood donors difficult for those who need transfusion and have developed anti-Jk3. This review describes methods used in identifying anti-Jk3 in four pregnant patients. Blood bank records were retrospectively reviewed to illustrate the common approach in anti-Jk3 identification. In all cases, pertinent blood bank history was gathered, and extended RBC phenotyping was performed, followed by adsorption studies and testing of selected RBCs. Underlying antibodies were found in two of the cases. This review also reiterates some common challenges encountered with Kidd antibody analysis and highlights the importance of patient ethnic ancestry and obtaining accurate patient transfusion history.
Topics: Antibody Specificity; Blood Banks; Blood Transfusion; Female; Humans; Kidd Blood-Group System; Pregnancy; Retrospective Studies
PubMed: 34170639
DOI: 10.21307/immunohematology-2021-013 -
Journal of Agricultural and Food... Feb 2024Flunixin (FLU) is a nonsteroidal drug that is widely used in animals, causing severe drug residues in animal-derived foods and environment. The development of...
Flunixin (FLU) is a nonsteroidal drug that is widely used in animals, causing severe drug residues in animal-derived foods and environment. The development of antibody-based rapid immunoassay methods is of great significance for the monitoring of FLU and its metabolite 5-hydroxyflunixin (5-FLU). We prepared monoclonal antibodies (mAbs) with different recognition spectra through FLU-keyhole limpet hemocyanin conjugates as immunogen coupled with antibody screening strategies. mAb5E6 and mAb6D7 recognized FLU with high affinity, and mAb2H5 and mAb4A4 recognized FLU and 5-FLU with broad specificity. Through evaluating the recognition of these mAbs against more than 11 structural analogues and employing computational chemistry, molecular docking, and molecular dynamics methodologies, we preliminarily determined the recognition epitope and recognition mechanism of these mAbs. Finally, an indirect competitive enzyme-linked immunosorbent assay for FLU based on mAb6D7 was developed, which exhibited limits of detection as low as 0.016-0.042 μg kg (L) in milk and muscle samples.
Topics: Animals; Antibody Formation; Molecular Docking Simulation; Immunoassay; Antibodies, Monoclonal; Enzyme-Linked Immunosorbent Assay; Antibody Specificity; Clonixin
PubMed: 38197248
DOI: 10.1021/acs.jafc.3c07766 -
Autoimmunity Reviews Dec 2021Alpha-enolase (Eno) is an ubiquitary glycolytic enzyme playing multiple functions that go well beyond its principal metabolic role of energy supplier during glycolysis.... (Review)
Review
Alpha-enolase (Eno) is an ubiquitary glycolytic enzyme playing multiple functions that go well beyond its principal metabolic role of energy supplier during glycolysis. Eno is localized in the cytoplasm, but also expressed on the cell membrane, where it binds plasminogen allowing its activation. Its shorter form, in the nucleus, acts as transcription factor. In inflammatory conditions, Eno undergoes post-translational modifications, such as citrullination, oxidation and phosphorylation. Eno is also an autoantigen in different disorders. In fact, autoantibodies to Eno have been detected in rheumatoid arthritis, lupus nephritis, primary glomerulonephritis, cancer, infections and other disorders, and in many cases they represent specific markers to be utilized in clinical practice. Anti-Eno antibodies in the different clinical conditions are not equal: they differ in isotype and often recognize different epitopes on the enzyme. IgG1 and IgG3 are prevalent in Rheumatoid Arthritis, IgG2 in Lupus nephritis and IgG4 in primary autoimmune glomerulopathy. This review analyzes the characteristics of anti-Eno autoantibodies in autoimmune disorders and cancer, describing their fine specificity and isotype restriction. The post-translational modifications that are target of autoantibodies are also discussed, as they represent the basis for elucidating the molecular mechanisms responsible for epitope generation. Despite an impressive amount of experimental work on anti-Eno antibodies, it is still necessary to validate the use of anti-Eno antibodies as biomarkers of selected diseases and extend the knowledge on the mechanisms of anti-Eno autoantibody production. Strategies that downmodulate the immune response to Eno may represent in the future novel approaches in the treatment of autoimmune disorders.
Topics: Antibody Specificity; Autoantibodies; Autoantigens; Humans; Lupus Erythematosus, Systemic; Lupus Nephritis; Phosphopyruvate Hydratase
PubMed: 34718161
DOI: 10.1016/j.autrev.2021.102977 -
Chembiochem : a European Journal of... Sep 2022Antibodies recognize their cognate antigens with high affinity and specificity, but the prediction of binding sites on the antigen (epitope) corresponding to a specific...
Antibodies recognize their cognate antigens with high affinity and specificity, but the prediction of binding sites on the antigen (epitope) corresponding to a specific antibody remains a challenging problem. To address this problem, we developed AbAdapt, a pipeline that integrates antibody and antigen structural modeling with rigid docking in order to derive antibody-antigen specific features for epitope prediction. In this study, we systematically assessed the impact of integrating the state-of-the-art protein modeling method AlphaFold with the AbAdapt pipeline. By incorporating more accurate antibody models, we observed improvement in docking, paratope prediction, and prediction of antibody-specific epitopes. We further applied AbAdapt-AF in an anti-receptor binding domain (RBD) antibody complex benchmark and found AbAdapt-AF outperformed three alternative docking methods. Also, AbAdapt-AF demonstrated higher epitope prediction accuracy than other tested epitope prediction tools in the anti-RBD antibody complex benchmark. We anticipate that AbAdapt-AF will facilitate prediction of antigen-antibody interactions in a wide range of applications.
Topics: Antibodies; Antibody Specificity; Antigens; Binding Sites, Antibody; Epitopes
PubMed: 35893479
DOI: 10.1002/cbic.202200303 -
MAbs 2020In the wake of the reproducibility crisis and numerous discussions on how commercially available antibodies as research tool contribute to it, The Antibody Society...
In the wake of the reproducibility crisis and numerous discussions on how commercially available antibodies as research tool contribute to it, The Antibody Society developed a series of 10 webinars to address the issues involved. The webinars were delivered by speakers with both academic and commercial backgrounds. This report highlights the problems, and offers solutions to help the scientific community appropriately identify the right antibodies and to validate them for their research and development projects. Despite the various solutions proposed here, they must be applied on a case-by-case basis. Each antibody must be verified based on the content of the product sheet, and subsequently through experimentation to confirm integrity, specificity and selectivity. Verification needs to focus on the precise application and tissue/cell type for which the antibody will be used, and all verification data must be reported openly. The various approaches discussed here all have caveats, so a combination of solutions must be considered.
Topics: Antibodies, Monoclonal; Antibody Specificity; Humans; Protein Engineering; Validation Studies as Topic
PubMed: 32748696
DOI: 10.1080/19420862.2020.1794421 -
Nature Communications Apr 2023During an infection the immune system produces pathogen-specific antibodies. These antibody repertoires become specific to the history of infections and represent a rich...
During an infection the immune system produces pathogen-specific antibodies. These antibody repertoires become specific to the history of infections and represent a rich source of diagnostic markers. However, the specificities of these antibodies are mostly unknown. Here, using high-density peptide arrays we examined the human antibody repertoires of Chagas disease patients. Chagas disease is a neglected disease caused by Trypanosoma cruzi, a protozoan parasite that evades immune mediated elimination and mounts long-lasting chronic infections. We describe a proteome-wide search for antigens, characterised their linear epitopes, and show their reactivity on 71 individuals from diverse human populations. Using single-residue mutagenesis we revealed the core functional residues for 232 of these epitopes. Finally, we show the diagnostic performance of identified antigens on challenging samples. These datasets enable the study of the Chagas antibody repertoire at an unprecedented depth and granularity, while also providing a rich source of serological biomarkers.
Topics: Humans; Trypanosoma cruzi; Epitopes; Antibody Specificity; Enzyme-Linked Immunosorbent Assay; Chagas Disease; Antigens, Protozoan; Antibodies; Americas; Antibodies, Protozoan
PubMed: 37012236
DOI: 10.1038/s41467-023-37522-9 -
Nature Communications May 2022The antibody response magnitude and kinetics may impact clinical severity, serological diagnosis and long-term protection of COVID-19, which may play a role in why...
The antibody response magnitude and kinetics may impact clinical severity, serological diagnosis and long-term protection of COVID-19, which may play a role in why children experience lower morbidity. We therefore tested samples from 122 children in Hong Kong with symptomatic (n = 78) and asymptomatic (n = 44) SARS-CoV-2 infections up to 200 days post infection, relative to 71 infected adults (symptomatic n = 61, and asymptomatic n = 10), and negative controls (n = 48). We assessed serum IgG antibodies to a 14-wide antigen panel of structural and accessory proteins by Luciferase Immuno-Precipitation System (LIPS) assay and circulating cytokines. Infected children have lower levels of Spike, Membrane, ORF3a, ORF7a, ORF7b antibodies, comparable ORF8 and elevated E-specific antibodies than adults. Combination of two unique antibody targets, ORF3d and ORF8, can accurately discriminate SARS-CoV-2 infection in children. Principal component analysis reveals distinct pediatric serological signatures, and the highest contribution to variance from adults are antibody responses to non-structural proteins ORF3d, NSP1, ORF3a and ORF8. From a diverse panel of cytokines that can modulate immune priming and relative inflammation, IL-8, MCP-1 and IL-6 correlate with the magnitude of pediatric antibody specificity and severity. Antibodies to SARS-CoV-2 internal proteins may become an important sero surveillance tool of infection with the roll-out of vaccines in the pediatric population.
Topics: Adult; Antibody Specificity; COVID-19; Child; Cytokines; Humans; Immunoglobulin G; SARS-CoV-2
PubMed: 35618731
DOI: 10.1038/s41467-022-30699-5 -
Immunobiology May 2022Previous studies have shown that polyreactive antibodies play an important role in the frontline defense against the dissemination of pathogens in the pre-immune host....
Previous studies have shown that polyreactive antibodies play an important role in the frontline defense against the dissemination of pathogens in the pre-immune host. Interestingly, antigen-binding polyreactivity can not only be inherent, but also acquired post-translationally. The ability of individual monoclonal IgG and IgE antibodies to acquire polyreactivity following contact with various agents that destabilize protein structure (urea, low pH) or have a pro-oxidative potential (heme, ferrous ions) has been studied in detail. However, to the best of our knowledge this property of human IgA has previously been described only cursorily. In the present study pooled human serum IgA and two human monoclonal IgA antibodies were exposed to buffers with acidic pH, to free heme or to ferrous ions, and the antigen-binding behavior of the native and modified IgA to viral and bacterial antigens were compared using immunoblot and ELISA. We observed a dose-dependent increase in reactivity to several bacterial extracts and to pure viral antigens. This newly described property of IgA may have therapeutic potential as has already been shown for pooled IgG with induced polyreactivity.
Topics: Antibodies, Monoclonal; Antibody Specificity; Heme; Humans; Immunoglobulin A; Immunoglobulin G; Ions
PubMed: 35429697
DOI: 10.1016/j.imbio.2022.152213 -
Methods in Molecular Biology (Clifton,... 2021Validation of antibody specificity is essential for the accurate evaluation of protein expression. For antibodies that recognize the gene products of the RAS family of...
Validation of antibody specificity is essential for the accurate evaluation of protein expression. For antibodies that recognize the gene products of the RAS family of oncogenes (HRAS, KRAS, and NRAS), an important challenge is the determination of selectivity for the four nearly identical HRAS, KRAS4A, KRAS4B, and NRAS proteins. With increasing appreciation for the distinct roles of the different RAS proteins in normal and neoplastic cells, there is a need for well-validated antibodies to evaluate the function and expression of the different RAS isoforms. Here we describe our experimental approaches to characterize RAS antibodies for their isoform- and mutant-specificity for use in immunoblot analyses.
Topics: Animals; Antibodies, Monoclonal; Antibody Specificity; Blotting, Western; Cells, Cultured; Embryo, Mammalian; Fibroblasts; Mice; Mutation; Protein Isoforms; ras Proteins
PubMed: 33977472
DOI: 10.1007/978-1-0716-1190-6_5 -
Papillomavirus Research (Amsterdam,... Dec 2019Human papillomavirus (HPV) infects and propagates in the cervical mucosal epithelium. Hence, in addition to assessing systemic immunity, the accurate measurement of... (Meta-Analysis)
Meta-Analysis Review
BACKGROUND
Human papillomavirus (HPV) infects and propagates in the cervical mucosal epithelium. Hence, in addition to assessing systemic immunity, the accurate measurement of cervical immunity is important to evaluate local immune responses to HPV infection and vaccination. This review discusses studies that investigated the presence of infection and vaccine-induced HPV-specific antibodies in cervicovaginal secretions (CVS).
METHODS
We searched the two main health sciences databases, PubMed and the ISI Web of Science, from the earliest dates available to March 2019. From the eligible publications, information was extracted regarding: (i) study design, (ii) the reported HPV-specific antibody concentrations in CVS (and the associated serum levels, when provided), (iii) the CVS collection method, and (iv) the immunoassays used.
RESULTS
The systematic search and selection process yielded 44 articles. The evidence of HPV-specific antibodies in CVS after natural infection (26/44) and HPV vaccination (18/44) is discussed. Many studies indicate that HPV-specific antibody detection in CVS is variable but feasible with a variety of collection methods and immunoassays. Most CVS samples were collected by cervicovaginal washing or wicks, and antibody presence was mostly determined by VLP-based ELISAs. The moderate to strong correlation between vaccine-induced antibody levels in serum and in CVS indicates that HPV vaccines generate antibodies that transudate through the cervical mucosal epithelium.
CONCLUSION
Although HPV-specific antibodies have lower titres in CVS than in serum samples, studies have shown that their detection in CVS is feasible. Nevertheless, the high variability of published observations and the lack of a strictly uniform, well-validated method for the collection, isolation and quantification of antibodies indicates a need for specific methods to improve and standardize the detection of HPV-specific antibodies in CVS.
Topics: Antibodies, Viral; Antibody Specificity; Cervix Uteri; Female; Host-Pathogen Interactions; Humans; Immunity, Mucosal; Papillomaviridae; Papillomavirus Infections; Papillomavirus Vaccines; Vaccination; Vagina
PubMed: 31494291
DOI: 10.1016/j.pvr.2019.100185