{"doi":"10.1101/2020.04.15.041996","title":"A facile method of mapping HIV-1 neutralizing epitopes using chemically masked cysteines and deep sequencing","abstract":"Abstract Identification of specific epitopes targeted by neutralizing antibodies is essential to advance epitope-based vaccine design strategies. We report a facile methodology for rapid epitope mapping of neutralizing antibodies (NAbs) against HIV-1 Envelope (Env) at single residue resolution, using Cys labeling, viral neutralization assays and deep sequencing. This was achieved by the generation of a library of Cys mutations in Env glycoprotein on the viral surface, covalent labeling of the Cys residues using a Cys reactive label that masks epitope residues, followed by infection of the labeled mutant virions in mammalian cells in the presence of NAbs. Env gene sequencing from NAb-resistant viruses was used to accurately delineate epitopes for the NAbs VRC01, PGT128 and PGT151. These agreed well with corresponding experimentally determined structural epitopes previously inferred from NAb:Env structures. HIV-1 infection is associated with complex and polyclonal antibody responses, typically composed of multiple antibody specificities. Deconvoluting the epitope specificities in a polyclonal response is a challenging task. We therefore extended our methodology to map multiple specificities of epitopes targeted in polyclonal sera, elicited in immunized animals as well as in an HIV-1 infected elite neutralizer capable of neutralizing Tier-3 pseudoviruses with high titers. The method can be readily extended to other viruses for which convenient reverse genetics or lentiviral surface display systems are available.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2020,"id":131566,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9559,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2020-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":424726,"name":"Rohan Roy Chowdhury","orcid":"0000-0003-3586-571X","position":1,"is_corresponding":false},{"id":424727,"name":"Kavyashree Manjunath","orcid":"0000-0002-1838-5362","position":2,"is_corresponding":false},{"id":326398,"name":"Luke Elizabeth Hanna","orcid":"0000-0002-6299-8211","position":3,"is_corresponding":false},{"id":424728,"name":"Raghavan Varadarajan","orcid":"0000-0002-0823-7577","position":4,"is_corresponding":false},{"id":424725,"name":"Rohini Datta","orcid":"0000-0003-2693-1922","position":0,"is_corresponding":true}],"reference_count":58,"raw_metadata":null,"created_at":"2026-07-18T23:16:03.875886Z","pmid":null,"pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}