Rhesus and cynomolgus macaque immunoglobulin heavy-chain genotyping yields comprehensive databases of germline VDJ alleles.

Definition of the specific germline immunoglobulin (Ig) alleles present in an individual is a critical first step to delineate the ontogeny and evolution of antigen-specific antibody responses. Rhesus and cynomolgus macaques are important animal models for pre-clinical studies, with four main sub-groups being used: Indian- and Chinese-origin rhesus macaques and Mauritian and Indonesian cynomolgus macaques. We applied the (Ig) gene inference tool IgDiscover and performed extensive Sanger sequencing-based genomic validation to define germline VDJ alleles in these 4 sub-groups, comprising 45 macaques in total.

Polyfunctional Tier 2-Neutralizing Antibodies Cloned following HIV-1 Env Macaque Immunization Mirror Native Antibodies in a Human Donor.

Vaccine efforts to combat HIV are challenged by the global diversity of viral strains and shielding of neutralization epitopes on the viral envelope glycoprotein trimer. Even so, the isolation of broadly neutralizing Abs from infected individuals suggests the potential for eliciting protective Abs through vaccination. This study reports a panel of 58 mAbs cloned from a rhesus macaque () immunized with envelope glycoprotein immunogens curated from an HIV-1 clade C-infected volunteer.

Vaccination with Glycan-Modified HIV NFL Envelope Trimer-Liposomes Elicits Broadly Neutralizing Antibodies to Multiple Sites of Vulnerability.

The elicitation of broadly neutralizing antibodies (bNAbs) against the HIV-1 envelope glycoprotein (Env) trimer remains a major vaccine challenge. Most cross-conserved protein determinants are occluded by self-N-glycan shielding, limiting B cell recognition of the underlying polypeptide surface. The exceptions to the contiguous glycan shield include the conserved receptor CD4 binding site (CD4bs) and glycoprotein (gp)41 elements proximal to the furin cleavage site.

Extensive dissemination and intraclonal maturation of HIV Env vaccine-induced B cell responses.

Well-ordered HIV-1 envelope glycoprotein (Env) trimers are prioritized for clinical evaluation, and there is a need for an improved understanding about how elicited B cell responses evolve following immunization. To accomplish this, we prime-boosted rhesus macaques with clade C NFL trimers and identified 180 unique Ab lineages from ∼1,000 single-sorted Env-specific memory B cells. We traced all lineages in high-throughput heavy chain (HC) repertoire (Rep-seq) data generated from multiple immune compartments and time points and expressed several as monoclonal Abs (mAbs).

High-Quality Library Preparation for NGS-Based Immunoglobulin Germline Gene Inference and Repertoire Expression Analysis.

Next generation sequencing (NGS) of immunoglobulin (Ig) repertoires (Rep-seq) enables examination of the adaptive immune system at an unprecedented level. Applications include studies of expressed repertoires, gene usage, somatic hypermutation levels, Ig lineage tracing and identification of genetic variation within the Ig loci through inference methods. All these applications require starting libraries that allow the generation of sequence data with low error rate and optimal representation of the expressed repertoire.

Particulate Array of Well-Ordered HIV Clade C Env Trimers Elicits Neutralizing Antibodies that Display a Unique V2 Cap Approach.

The development of soluble envelope glycoprotein (Env) mimetics displaying ordered trimeric symmetry has ushered in a new era in HIV-1 vaccination. The recently reported native, flexibly linked (NFL) design allows the generation of native-like trimers from clinical isolates at high yields and homogeneity. As the majority of infections world-wide are of the clade C subtype, we examined responses in non-human primates to well-ordered subtype C 16055 trimers administered in soluble or high-density liposomal formats.

Production of individualized V gene databases reveals high levels of immunoglobulin genetic diversity.

Comprehensive knowledge of immunoglobulin genetics is required to advance our understanding of B cell biology. Validated immunoglobulin variable (V) gene databases are close to completion only for human and mouse. We present a novel computational approach, IgDiscover, that identifies germline V genes from expressed repertoires to a specificity of 100%.

Antibodies Elicited by Multiple Envelope Glycoprotein Immunogens in Primates Neutralize Primary Human Immunodeficiency Viruses (HIV-1) Sensitized by CD4-Mimetic Compounds.

Unlabelled: The human immunodeficiency virus (HIV-1) envelope glycoproteins (Env) mediate virus entry through a series of complex conformational changes triggered by binding to the receptors CD4 and CCR5/CXCR4. Broadly neutralizing antibodies that recognize conserved Env epitopes are thought to be an important component of a protective immune response. However, to date, HIV-1 Env immunogens that elicit broadly neutralizing antibodies have not been identified, creating hurdles for vaccine development.

Diverse antibody genetic and recognition properties revealed following HIV-1 envelope glycoprotein immunization.

Isolation of mAbs elicited by vaccination provides opportunities to define the development of effective immunity. Ab responses elicited by current HIV-1 envelope glycoprotein (Env) immunogens display narrow neutralizing activity with limited capacity to block infection by tier 2 viruses. Intense work in the field suggests that improved Env immunogens are forthcoming, and it is therefore important to concurrently develop approaches to investigate the quality of vaccine-elicited responses at a higher level of resolution.