{"doi":"10.1016/j.xpro.2024.103509","title":"Protocol to study secretome interactions using extracellular proximity labeling","abstract":"Biotin ligase-based proximity ligation is a widely used, highly effective technique for the study of in vivo protein-protein interactions. However, there are few reports and little consensus on the most effective methods for studying the proximal interactomes of secreted factors. Here, we present a protocol for studying extracellular proximal interactomes using an adaptation of TurboID/BioID2-based proximity ligation. We describe steps for cell preparation, sample collection, and initial processing. We then detail procedures for biotinylated protein enrichment, on-bead digestion, and post-pull-down processing. For complete details on the use and execution of this protocol, please refer to Peeney et al. 1 • Guidance on the production of secreted biotin ligase fusion proteins • Steps for initiating extracellular proximity labeling experiments • Instructions for processing unlabeled and isobaric labeled samples • Best practices for data processing and interpretation Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Biotin ligase-based proximity ligation is a widely used, highly effective technique for the study of in vivo protein-protein interactions. However, there are few reports and little consensus on the most effective methods for studying the proximal interactomes of secreted factors. Here, we present a protocol for studying extracellular proximal interactomes using an adaptation of TurboID/BioID2-based proximity ligation. We describe steps for cell preparation, sample collection, and initial processing. We then detail procedures for biotinylated protein enrichment, on-bead digestion, and post-pull-down processing.","journal":"STAR Protocols","year":2024,"id":470111,"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":3,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9595,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":882602,"name":"Sadeechya Gurung","orcid":null,"position":1,"is_corresponding":false},{"id":1058162,"name":"Sasha Coates-Park","orcid":"0009-0005-2291-5337","position":2,"is_corresponding":false},{"id":400996,"name":"Yueqin Liu","orcid":null,"position":3,"is_corresponding":false},{"id":1304597,"name":"Anshika Govil","orcid":null,"position":4,"is_corresponding":false},{"id":881695,"name":"William G. Stetler‐Stevenson","orcid":"0000-0002-5500-5808","position":5,"is_corresponding":false},{"id":881693,"name":"David Peeney","orcid":"0000-0002-9126-9923","position":6,"is_corresponding":false},{"id":1170858,"name":"Joshua A. Rich","orcid":"0000-0002-2206-5214","position":0,"is_corresponding":true}],"reference_count":10,"raw_metadata":null,"created_at":"2026-07-19T02:05:36.656771Z","pmid":"39673706","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":[]}