Identification and Characterization of Unique Neutralizing Antibodies to Mouse EGF Receptor.
Ontology highlight
ABSTRACT:
SUBMITTER: Jae Huh W
PROVIDER: S-EPMC7103561 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:This SuperSeries is composed of the SubSeries listed below.
2024-06-04 | GSE229123 | GEO
Project description:Exosomes are small, 40-130 nm secreted extracellular vesicles that recently have become the subject of intense focus as agents of intercellular communication, disease biomarkers and potential vehicles for drug delivery. It is currently unknown whether a cell produces different populations of exosomes with distinct cargo and separable functions. To address this question, high-resolution methods are needed. Using a commercial flow cytometer and directly labelled fluorescent antibodies, we show the feasibility of using fluorescence-activated vesicle sorting (FAVS) to analyse and sort individual exosomes isolated by sequential ultracentrifugation from the conditioned medium of DiFi cells, a human colorectal cancer cell line. EGFR and the exosomal marker, CD9, were detected on individual DiFi exosomes by FAVS; moreover, both markers were identified by high-resolution stochastic optical reconstruction microscopy on individual, approximately 100 nm vesicles from flow-sorted EGFR/CD9 double-positive exosomes. We present evidence that the activation state of EGFR can be assessed in DiFi-derived exosomes using a monoclonal antibody (mAb) that recognizes "conformationally active" EGFR (mAb 806). Using human antigen-specific antibodies, FAVS was able to detect human EGFR and CD9 on exosomes isolated from the plasma of athymic nude mice bearing DiFi tumour xenografts. Multicolour FAVS was used to simultaneously identify CD9, EGFR and an EGFR ligand, amphiregulin (AREG), on human plasma-derived exosomes from 3 normal individuals. These studies demonstrate the feasibility of FAVS to both analyse and sort individual exosomes based on specific cell-surface markers. We propose that FAVS may be a useful tool to monitor EGFR and AREG in circulating exosomes from individuals with colorectal cancer and possibly other solid tumours.
| S-EPMC4921784 | biostudies-literature
Project description:Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract infection in young children and older adults. Currently, no licensed vaccine is available, and therapeutic options are limited. The primary target of neutralizing antibodies to RSV is the surface fusion (F) glycoprotein. Understanding the recognition of antibodies with high neutralization potencies to RSV F antigen will provide critical insights in developing efficacious RSV antibodies and vaccines. In this study, we isolated and characterized a panel of monoclonal antibodies (mAbs) with high binding affinity to RSV prefusion F trimer and neutralization potency to RSV viruses. The mAbs were mapped to previously defined antigenic sites, and some that mapped to the same antigenic sites showed remarkable diversity in specificity, binding, and neutralization potencies. We found that the isolated site III mAbs shared highly conserved germline V-gene usage, but had different cross-reactivities to human metapneumovirus (hMPV), possibly due to the distinct modes/angles of interaction with RSV and hMPV F proteins. Furthermore, we identified a subset of potent RSV/hMPV cross-neutralizing mAbs that target antigenic site IV and the recently defined antigenic site V, while the majority of the mAbs targeting these two sites only neutralize RSV. Additionally, the isolated mAbs targeting site Ø were mono-specific for RSV and showed a wide range of neutralizing potencies on different RSV subtypes. Our data exemplify the diversity of anti-RSV mAbs and provide new insights into the immune recognition of respiratory viruses in the Pneumoviridae family.
| S-EPMC6816417 | biostudies-literature
Project description:We generated the humoral immune repertoire of circulating memory B cells from healthy and HIV infected immune donors. These repertoires were used to predict specific HIV antibodies directed against gp120/gp41.
2024-06-04 | GSE229122 | GEO
Project description:We generated the humoral immune repertoire of circulating memory B cells from healthy and HIV infected immune donors. These repertoires were used to predict specific HIV antibodies directed against gp120/gp41.
2024-06-04 | GSE228912 | GEO
Project description:Abs that stimulate the thyrotropin receptor (TSHR), the cause of Graves' hyperthyroidism, only develop in humans. TSHR Abs can be induced in mice by immunization, but studying pathogenesis and therapeutic intervention requires a model without immunization. Spontaneous, iodine-accelerated, thyroid autoimmunity develops in NOD.H2(h4) mice associated with thyroglobulin and thyroid-peroxidase, but not TSHR, Abs. We hypothesized that transferring the human TSHR A-subunit to NOD.H2(h4) mice would result in loss of tolerance to this protein. BALB/c human TSHR A-subunit mice were bred to NOD.H2(h4) mice, and transgenic offspring were repeatedly backcrossed to NOD.H2(h4) mice. All offspring developed Abs to thyroglobulin and thyroid-peroxidase. However, only TSHR-transgenic NOD.H2(h4) mice (TSHR/NOD.H2(h4)) developed pathogenic TSHR Abs as detected using clinical Graves' disease assays. As in humans, TSHR/NOD.H2(h4) female mice were more prone than male mice to developing pathogenic TSHR Abs. Fortunately, in view of the confounding effect of excess thyroid hormone on immune responses, spontaneously arising pathogenic human TSHR Abs cross-react poorly with the mouse TSHR and do not cause thyrotoxicosis. In summary, the TSHR/NOD.H2(h4) mouse strain develops spontaneous, iodine-accelerated, pathogenic TSHR Abs in female mice, providing a unique model to investigate disease pathogenesis and test novel TSHR Ag-specific immunotherapies aimed at curing Graves' disease in humans.
| S-EPMC4402262 | biostudies-literature
Project description:We generated the humoral immune repertoire of circulating memory B cells from healthy and HIV infected immune donors. These repertoires were used to predict specific HIV antibodies directed against gp120/gp41.
2024-06-04 | GSE229121 | GEO
Project description:A less than adequate therapeutic plan for the treatment of anthrax in the 2001 bioterrorism attacks has highlighted the importance of developing alternative or complementary therapeutic approaches for biothreat agents. In these regards passive immunization possesses several important advantages over active vaccination and the use of antibiotics, as it can provide immediate protection against Bacillus anthracis. Herein, we report the selection and characterization of several human monoclonal neutralizing antibodies against the toxin of B. anthracis from a phage displayed human scFv library. In total 15 clones were selected with distinct sequences and high specificity to protective antigen and thus were the subject of a series of both biophysical and cell-based cytotoxicity assays. From this panel of antibodies a set of neutralizing antibodies were identified, of which clone A8 recognizes the lethal (and/or edema) factor binding domain, and clones F1, G11, and G12 recognize the cellular receptor binding domain found within the protective antigen. It was noted that all clones distinguish a conformational epitope existing on the protective antigen; this steric relationship was uncovered using a sequential epitope mapping approach. For each neutralizing antibody, the kinetic constants were determined by surface plasmon resonance, while the potency of protection was established using a two-tier macrophage cytotoxicity assay. Among the neutralizing antibodies identified, clone F1 possessed the highest affinity to protective antigen, and provided superior protection from lethal toxin in the cell cytotoxicity assay. The data presented provide the ever-growing arsenal of immunological and functional analysis of monoclonal antibodies to the exotoxins of anthrax. In addition it grants new candidates for the prophylaxis and therapeutic treatment against this toxin.
| S-EPMC2268906 | biostudies-literature
Project description:Many human monoclonal antibodies that neutralize multiple clades of HIV-1 are polyreactive and bind avidly to mammalian autoantigens. Indeed, the generation of neutralizing antibodies to the 2F5 and 4E10 epitopes of HIV-1 gp41 in man may be proscribed by immune tolerance since mice expressing the VH and VL regions of 2F5 have a block in B-cell development characteristic of central tolerance. This developmental blockade implies the presence of tolerizing autoantigens that are mimicked by the membrane-proximal external region of HIV-1 gp41. Here we identify human kynureninase (KYNU) and splicing factor 3b subunit 3 (SF3B3) as the primary conserved, vertebrate self-antigens recognized by the 2F5 and 4E10 antibodies, respectively. 2F5 binds the H4 domain of KYNU which contains the complete 2F5 linear epitope (ELDKWA). 4E10 recognizes a conformational epitope of SF3B3 that is strongly dependent on hydrophobic interactions. Opossums carry a rare KYNU H4 domain that abolishes 2F5 binding, but retain all SF3B3 4E10 epitopes. Immunization of opossums with HIV-1 gp140 induced extraordinary titers of serum antibody to the 2F5 ELDKWA epitope but little or nothing to the 4E10 determinant. Identification of structural motif shared by vertebrates and HIV-1 provides direct evidence that immunological tolerance can impair humoral responses to HIV-1. The invitrogen protoarray that contains >9,400 recombinant human proteins was used to identify self-ligands that are recognized by broadly neutralizing HIV-1 antibodies 2F5 and 4E10. An isotype-matched human myeloma protein (151K, Southern Biotech) was used as control.
2012-12-03 | E-GEOD-42673 | biostudies-arrayexpress
Project description:Many human monoclonal antibodies that neutralize multiple clades of HIV-1 are polyreactive and bind avidly to mammalian autoantigens. Indeed, the generation of neutralizing antibodies to the 2F5 and 4E10 epitopes of HIV-1 gp41 in man may be proscribed by immune tolerance since mice expressing the VH and VL regions of 2F5 have a block in B-cell development characteristic of central tolerance. This developmental blockade implies the presence of tolerizing autoantigens that are mimicked by the membrane-proximal external region of HIV-1 gp41. Here we identify human kynureninase (KYNU) and splicing factor 3b subunit 3 (SF3B3) as the primary conserved, vertebrate self-antigens recognized by the 2F5 and 4E10 antibodies, respectively. 2F5 binds the H4 domain of KYNU which contains the complete 2F5 linear epitope (ELDKWA). 4E10 recognizes a conformational epitope of SF3B3 that is strongly dependent on hydrophobic interactions. Opossums carry a rare KYNU H4 domain that abolishes 2F5 binding, but retain all SF3B3 4E10 epitopes. Immunization of opossums with HIV-1 gp140 induced extraordinary titers of serum antibody to the 2F5 ELDKWA epitope but little or nothing to the 4E10 determinant. Identification of structural motif shared by vertebrates and HIV-1 provides direct evidence that immunological tolerance can impair humoral responses to HIV-1.
2012-12-03 | GSE42673 | GEO