PO.CH02.02 · 化学
用于安全、可扩展和灵活的单细胞多组学研究的无甲醛固定
Formaldehyde-free fixation for safe, scalable and flexible single-cell multiomics studies
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摘要 Abstract
中文摘要
灵活的单细胞多组学方法的开发和商业化，使得研究者能够选择和实施为其感兴趣的疾病模型提供所需生物学深度和背景的检测。多组学方法通常在使用新鲜、高质量样本时获得最佳结果，以高效恢复天然生物学状态。然而，这些动态样本给较大规模的临床研究带来了物流挑战，限制了研究范围和可重复性。甲醛（PFA）固定一直是组织保存的标准，因其可及性和稳定性能够简化临床样本制备以及来自大型患者队列的生物样本库建设。然而，PFA的应用早于人们对其致癌性和核酸交联作用的普遍认知，这限制了未来的转录组和基因组研究。与组织不同，单细胞悬液的固定尚未标准化到这一程度，这为寻求更安全、更灵活的样本保存方案提供了宝贵的机会。为此，我们评估了因相较PFA具有上述优势而日益受到研究的固定剂，目标是提出一种可供进一步研究的替代性单细胞固定剂方案。我们使用各种细胞和分子分析工具优化了每种替代固定方案，表明其具有广泛的多组学和临床兼容性。最佳配方用于处理外周血单核细胞（PBMCs）及其细胞核，并通过遗传（ATAC-seq）、转录组（WTA）和蛋白质组（CITE-seq）检测进行多组学评估，与活细胞和PFA对照进行比较。我们的数据鉴定出一种安全稳定的新型固定剂配方，可在模拟生物样本库建设过程中保存PBMC样本，并提供了与PFA相当或超越PFA的广泛多组学兼容性。这种替代方案及其新颖的制备方法，在学术界和工业界关于单细胞固定方案的研究中尚未得到充分体现，而这正值大型队列研究的关键时期。随着这些大型单细胞图谱和临床研究日益普遍，需要对这种更安全的替代方案进行进一步的开发、优化和验证。此类研究的数据正在塑造该领域基础性的新型虚拟细胞模型和临床标准操作规程，因此有必要进一步审视具有更灵活检测兼容性的更安全替代方案。仅供研究使用。不用于诊断或治疗程序。©2025 BD。保留所有权利。NPM-7460 (v1.0) 1125
查看英文原文 English abstract
The development and commercialization of flexible single-cell multiomics methods has allowed for investigator selection and implementation of assays which provide the desired biological depth and context for their disease model of interest. Multiomic methods generally achieve best results with fresh, high-quality samples for efficient recovery of the native biological state. These dynamic samples, however, pose logistical challenges for larger clinical studies, limiting scope and reproducibility. Formaldehyde (PFA) fixation has been the standard for tissue preservation, due to its accessibility and stability enabling simplified clinical sample preparation and biobanking from large patient cohorts. The implementation of PFA, however, predates common knowledge of its carcinogenicity and nucleic acid crosslinking, limiting future transcriptomic and genomic study. Unlike tissue, fixation of single cell suspensions has not yet been standardized to this extent, offering a valuable opportunity to survey more safe and flexible solutions for sample preservation. To address this, we evaluated fixatives that have increasingly been investigated for these advantages over PFA with the goal of suggesting an alternative single-cell fixative solution for further investigation. We optimized each alternative fixation solution using various cellular and molecular analysis tools suggesting broad multiomic and clinical compatibility. Optimal formulations were used to treat peripheral blood monocytes (PBMCs) and their nuclei and analyzed by genetic (ATAC-seq), transcriptomic (WTA) and proteomic (CITE-seq) assays for multiomic evaluation against live and PFA controls. Our data identified a safe and stable novel fixative formulation for preserving PBMC samples throughout a mock biobanking process and gave broad multiomic compatibility comparable or exceeding that of PFA.This alternative and its novel preparation are underrepresented in academic and industry studies on single-cell fixation solutions at a critical time for large cohort studies. Further development, optimization and demonstration of this safer alternative is needed as these large single cell atlas and clinical studies become increasingly common. Such studies' data are shaping new virtual cell models and clinical standard operating procedures foundational to this field and necessitate further review of safer alternatives with more flexible assay compatibility.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.©2025 BD. All rights reserved. NPM-7460 (v1.0) 1125
利益披露 Disclosure
J. Olives, None..
H. Huang, None..
Z. Zhang, None..
L. Chen, None..
R. Nguyen, None..
H. Song, None..
A. Ayer, None.