{"doi":"10.1002/hon.70096_506","title":"506 | EVALUATION OF ULTRASENSITIVE IN SITU HYBRIDIZATION MRNA TECHNOLOGY VERSUS IMMUNOHISTOCHEMISTRY FOR DETERMINING LIGHT CHAIN RESTRICTION IN B‐CELL NEOPLASMS","abstract":"Introduction: Determination of immunoglobulin light chain restriction status is crucial in determining clonality in suspected B-cell malignancies. Conventional immunohistochemistry (IHC) and in-situ hybridization (ISH) assays are useful for determining clonality in plasma cell tumors but fail to detect surface immunoglobulins expression status in most B-cell neoplasms. Ultrasensitive dual ISH mRNA is an emerging technology that detects a broader spectrum of B-cell non-Hodgkin lymphomas including cases without plasmacytic differentiation. In this study, staining characteristics and performance of two ultrasensitive ISH and one IHC assay for light chain restriction were assessed. Methods: 200 bone marrow and lymphoid tissue specimens with known historical light chain restriction results that met inclusion and exclusion criteria were identified at two study sites. 160 cases were either B-cell lymphomas or plasma cell malignancies, while 40 cases were reactive mimics. All cases were stained with the VENTANA Kappa and Lambda Dual ISH mRNA Probe Cocktail, Kappa and lambda RNAScope ISH Probes, and Dako kappa and lambda light chain IHC antibodies on a BenchMark ULTRA or DISCOVERY ULTRA instrument and independently interpreted for restriction status by four study pathologists across two study sites using glass slides or digital images. The pathologists' assessments for each case were compared to the available historical flow derived kappa and lambda (K/L) restriction status and final pathology diagnosis associated with the same patient to assess the assay performance. Results: All four pathologists subjectively evaluated each case by categorizing as either easy, moderate, or difficult. With both glass and digital slides, 70.6%, 61.6%, and 38.8% were rated as easy to interpret by the four pathologists with the ultrasensitive VENTANA Kappa and Lambda Dual ISH mRNA Probe Cocktail, ultrasensitive RNAScope ISH, and Dako IHC assays, respectively. Agreement with all three testing methods with historical flow cytometry was performed. Overall percent agreement with the VENTANA assay was 92.7%, with the RNAScope assay was 92.3%, and with the Dako assay was 72.9% (Table 1). Conclusions: Slides stained with either ultrasensitive kappa and lambda ISH for light chain restriction were subjectively easier to interpret compared to kappa and lambda immunohistochemistry. Using kappa and lambda light chain restriction by flow cytometry as a gold standard, both methods of ultrasensitive ISH had similar concordance but were both superior to immunohistochemistry. Ultrasensitive ISH for light chain restriction is a viable option for pathologists to determine clonality for lymphoid and bone marrow specimens for B-cell neoplasms. BENCHMARK, DISCOVERY, and VENTANA are trademarks of Roche. Research funding declaration: Study funded by Roche Diagnostics Solutions Keywords: non-Hodgkin; pathology and classification of lymphomas; diagnostic and prognostic biomarkers Potential sources of conflict of interest: B. Su Employment or leadership position: Roche Stock ownership: Roche M. Canamero Employment or leadership position: Roche S. Gladden Employment or leadership position: Roche B. Mosteller Employment or leadership position: Roche S. Rashidi Employment or leadership position: Roche J. Su Employment or leadership position: Roche","journal":"Hematological Oncology","year":2025,"id":568504,"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":0.9626,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1472763,"name":"Marta Cañamero","orcid":"0000-0003-2239-4285","position":1,"is_corresponding":false},{"id":1473324,"name":"Sarah Gladden","orcid":null,"position":2,"is_corresponding":false},{"id":1473325,"name":"Rebecca L. Kunak","orcid":null,"position":3,"is_corresponding":false},{"id":1473326,"name":"Brian Mosteller","orcid":null,"position":4,"is_corresponding":false},{"id":1473327,"name":"S. Rashidi","orcid":null,"position":5,"is_corresponding":false},{"id":1472764,"name":"Jens Redemann","orcid":"0000-0002-2404-7984","position":6,"is_corresponding":false},{"id":839357,"name":"Juan Su","orcid":"0000-0001-6239-1711","position":7,"is_corresponding":false},{"id":1473328,"name":"M A Vasef","orcid":null,"position":8,"is_corresponding":false},{"id":1472765,"name":"Ivonne Vázquez","orcid":"0000-0002-5305-1560","position":9,"is_corresponding":false},{"id":1472766,"name":"Luís Colomo","orcid":"0000-0001-5236-5085","position":10,"is_corresponding":false},{"id":272041,"name":"Bing Su","orcid":"0000-0002-4379-9014","position":0,"is_corresponding":true}],"reference_count":0,"raw_metadata":null,"created_at":"2026-07-19T02:56:52.212268Z","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":[]}