The Experts below are selected from a list of 17205 Experts worldwide ranked by ideXlab platform

Aleksandar Antanasijevic - One of the best experts on this subject based on the ideXlab platform.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    Nature Communications, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    bioRxiv, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    In Brief Herein, we evaluated the immunogenicity of several BG505 SOSIP-based HIV Env immunogens in the rhesus macaque animal model using a combination of serology and biophysical approaches. We applied electron cryo-microscopy for high-resolution mapping of elicited polyclonal Antibody responses, which provided detailed insights into the binding modes of the most common classes of antibodies elicited by BG505 SOSIP immunogens as well as the critical differences in immunogenicity that can occur as a consequence of engineered stabilizing mutations and partial glycan occupancy at different sites. Summary Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic. Graphical abstract

Jennifer B Silverman - One of the best experts on this subject based on the ideXlab platform.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    Nature Communications, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    bioRxiv, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    In Brief Herein, we evaluated the immunogenicity of several BG505 SOSIP-based HIV Env immunogens in the rhesus macaque animal model using a combination of serology and biophysical approaches. We applied electron cryo-microscopy for high-resolution mapping of elicited polyclonal Antibody responses, which provided detailed insights into the binding modes of the most common classes of antibodies elicited by BG505 SOSIP immunogens as well as the critical differences in immunogenicity that can occur as a consequence of engineered stabilizing mutations and partial glycan occupancy at different sites. Summary Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic. Graphical abstract

Leigh M Sewall - One of the best experts on this subject based on the ideXlab platform.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    Nature Communications, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    bioRxiv, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    In Brief Herein, we evaluated the immunogenicity of several BG505 SOSIP-based HIV Env immunogens in the rhesus macaque animal model using a combination of serology and biophysical approaches. We applied electron cryo-microscopy for high-resolution mapping of elicited polyclonal Antibody responses, which provided detailed insights into the binding modes of the most common classes of antibodies elicited by BG505 SOSIP immunogens as well as the critical differences in immunogenicity that can occur as a consequence of engineered stabilizing mutations and partial glycan occupancy at different sites. Summary Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic. Graphical abstract

Christopher A Cottrell - One of the best experts on this subject based on the ideXlab platform.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    Nature Communications, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic.

  • polyclonal Antibody responses to hiv env immunogens resolved using cryoem
    bioRxiv, 2021
    Co-Authors: Aleksandar Antanasijevic, Leigh M Sewall, Christopher A Cottrell, Diane G Carnathan, Luis E Jimenez, Julia T Ngo, Jennifer B Silverman
    Abstract:

    In Brief Herein, we evaluated the immunogenicity of several BG505 SOSIP-based HIV Env immunogens in the rhesus macaque animal model using a combination of serology and biophysical approaches. We applied electron cryo-microscopy for high-resolution mapping of elicited polyclonal Antibody responses, which provided detailed insights into the binding modes of the most common classes of antibodies elicited by BG505 SOSIP immunogens as well as the critical differences in immunogenicity that can occur as a consequence of engineered stabilizing mutations and partial glycan occupancy at different sites. Summary Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal Antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal Antibody responses without the need for monoclonal Antibody Isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic. Graphical abstract

Itai Benhar - One of the best experts on this subject based on the ideXlab platform.

  • Selection of antibodies from synthetic Antibody libraries.
    Archives of Biochemistry and Biophysics, 2012
    Co-Authors: Noa Harel Inbar, Itai Benhar
    Abstract:

    Abstract More than 2 dozen years had passed since the field of Antibody engineering was established, with the first reports of bacterial [1] , [2] , [3] and mammalian cells [4] expression of recombinant Antibody fragments, and in that time a lot of effort was dedicated to the development of efficient technological means, intended to assist in the creation of therapeutic monoclonal antibodies (mAbs). Research focus was given to two intertwined technological aspects: the selection platform and the recombinant Antibody repertoires. In accordance with these areas of interest, it is the goal of this chapter to describe the various selection tools and Antibody libraries existing, with emphasis on the later, and their applications. This chapter gives a far from exhaustive, subjective “historic account” of the field, describing the selection platforms, the different formats of Antibody repertoires and the applications of both for selecting recombinant antibodies. Several excellent books provide detailed protocols for constructing Antibody libraries and selecting antibodies from those libraries [5] , [6] , [7] , [8] , [9] , [10] , [11] , [12] , [13] . Such books may guide a newcomer to the field in the fine details of Antibody engineering. We would like to offer advice to the novice: although seemingly simple, effective library construction and Antibody Isolation provide best benefits in the hands of professionals. It is an art as much as it is science.

  • Antibody Isolation from immunized animals: Comparison of phage display and Antibody discovery via v gene repertoire mining
    Protein Engineering Design and Selection, 2012
    Co-Authors: Ido Saggy, Leeron Shefet-carasso, Limor Nahary, George Georgiou, Yariv Wine, Itai Benhar
    Abstract:

    Phage display has enabled the rapid Isolation of antigen-specific antibodies from combinatorial libraries of V(H) and V(L) genes obtained from lymphocytes of immunized animals. Recently, a different approach to Antibody Isolation that circumvents library screening and instead relies on the mining of the V(H) and V(L) gene repertoires obtained by high throughput sequencing of cDNAs from bone marrow Antibody-secreting cells was reported. Here we compared the antibodies obtained via phage library screening or via repertoire mining of V gene cDNAs obtained from total splenocytes of mice immunized with the hapten trinitrophenyl (TNP) conjugated to carrier proteins. We show that, despite the large heterogeneity of B lymphocytes in the spleen, the most abundant V genes encoded antigen-specific antibodies, indicating that total splenocytes can be used in place of bone marrow plasma cells for Antibody discovery at least in high titer animals. While both phage display and repertoire mining yielded antigen-specific antibodies showing comparable affinities by enzyme-linked immunosorbent assay analysis, clones obtained by the latter approach displayed higher selectivity towards TNP relative to control haptens. Interestingly, the Antibody genes isolated by phage display were of low abundance or absent from the V gene repertoire obtained by 454 sequencing. Similarly, the highly abundant V genes identified by repertoire mining, that as soluble antibodies were antigen-specific, were found to be poorly displayed on phage and were not enriched by phage panning. Thus, our results reveal that phage display and repertoire mining of immune repertoires are complementary technologies that can yield different antigen-specific Antibody clones.