Abstract 1434: Advancing multiomic single-cell ATAC-seq assays by expanding scalability and flexibility using nuclear sample multiplexing and gentle sample preservation techniques
Authors: Hongduan Huang, Quyen Bao, Rosary Nguyen, Hye-Won Song, Aruna Ayer
Journal: Cancer Research (2025)

Abstract Recent advances in multiomic single-cell ATAC-seq assay development have enabled researchers to unravel complex biological questions by simultaneously profiling chromatin accessibility and gene expression in a single nucleus. One major challenge, however, remains in boosting scalability and flexibility while still capturing high-quality multiomic data. Sample preparation and batching could be the key to resolving these issues. Here we demonstrate the feasibility of sample multiplexing utilizing oligo-conjugated nucleoporin antibody for nuclei hashing, allowing higher throughput and more efficient workflows. Up to 6 nuclei samples can be tagged separately and then pooled as one multiplex sample in a single ATAC-seq experiment, which minimizes batch effects and lowers per-sample cost. Additionally, we show the compatibility of sample preservation prior to scATAC-seq workflows using BD® OMICS-Guard Sample Preservation Buffer, providing added flexibility of timing without compromising the health of the cell. Moreover, we have combined these approaches and tested sample multiplexing capabilities on stored samples, further enhancing assays’ scalability and flexibility. Our data showed that PBMCs preserved with BD® OMICS-Guard Buffer consistently provided high sensitivity and high specificity ATAC-seq data. High ATAC peak correlation and whole transcriptome (WTA) gene expression correlation indicated that chromatin accessibility, RNA integrity and cell biology were well preserved by BD® OMICS-Guard Buffer. Successful sample multiplexing was observed in both cell line samples and PBMCs. Clear sample de-multiplexing was achieved with &amp;gt; 95% cell identification efficacy and &amp;gt; 95% tagging specificity. No significant impact on scATAC-seq or scRNA-seq data quality was observed, compared to unstained controls. Finally, to understand the usability of both techniques, we tested nuclei tagging of nuclei isolated from cells preserved with OMICS-Guard. The data showed that nuclei tagging and de-multiplexing efficiency remained unaffected by cell preservation. Together, our data support that the applications of BD® OMICS-Guard Sample Preservation Buffer and/or nuclei tagging increase scalability and flexibility to design and perform multiomic ATAC-seq and RNA-seq experiments without compromising data quality, which can profoundly impact experiment costs and overall success. For Research-Use Only. Not for use in diagnostic or therapeutic procedures. BD and the BD-Logo are trademarks of Becton, Dickinson and Company. © 2024 BD. All rights reserved. NPM-5011 (v1.0) 1024 Citation Format: Hongduan Huang, Quyen Bao, Rosary Nguyen, Hye-Won Song, Aruna Ayer. Advancing multiomic single-cell ATAC-seq assays by expanding scalability and flexibility using nuclear sample multiplexing and gentle sample preservation techniques [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1434.