Multiplex Protein Imaging through PACIFIC: Photoactive Immunofluorescence with Iterative Cleavage

IF 3.8 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY

ACS Bio & Med Chem Au Pub Date : 2023-04-28 DOI:10.1021/acsbiomedchemau.3c00018

Fei Ji, Moises Hur, Sungwon Hur, Siwen Wang, Priyanka Sarkar, Shiqun Shao, Desiree Aispuro, Xu Cong, Yanhao Hu, Zhonghan Li* and Min Xue*,

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引用次数: 0

Abstract

Multiplex protein imaging technologies enable deep phenotyping and provide rich spatial information about biological samples. Existing methods have shown great success but also harbored trade-offs between various pros and cons, underscoring the persisting necessity to expand the imaging toolkits. Here we present PACIFIC: photoactive immunofluorescence with iterative cleavage, a new modality of multiplex protein imaging methods. PACIFIC achieves iterative multiplexing by implementing photocleavable fluorophores for antibody labeling with one-step spin-column purification. PACIFIC requires no specialized instrument, no DNA encoding, or chemical treatments. We demonstrate that PACIFIC can resolve cellular heterogeneity in both formalin-fixed paraffin-embedded (FFPE) samples and fixed cells. To further highlight how PACIFIC assists discovery, we integrate PACIFIC with live-cell tracking and identify phosphor-p70S6K as a critical driver that governs U87 cell mobility. Considering the cost, flexibility, and compatibility, we foresee that PACIFIC can confer deep phenotyping capabilities to anyone with access to traditional immunofluorescence platforms.

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多重蛋白成像通过太平洋:光活性免疫荧光与迭代切割

多重蛋白质成像技术能够进行深入的表型分析，并提供有关生物样本的丰富空间信息。现有的方法取得了巨大的成功，但也在各种利弊之间进行了权衡，强调了扩展成像工具包的持续必要性。在这里，我们介绍了太平洋：迭代切割的光活性免疫荧光，一种新的多重蛋白质成像方法。PACIFIC通过一步旋转柱纯化实现抗体标记的可光裂解荧光团，实现迭代复用。PACIFIC不需要专门的仪器，不需要DNA编码或化学处理。我们证明PACIFIC可以解决福尔马林固定石蜡包埋（FFPE）样品和固定细胞中的细胞异质性。为了进一步强调PACIFIC如何帮助发现，我们将PACIFIC与活细胞跟踪相结合，并将磷酸-p70S6K确定为控制U87细胞迁移的关键驱动因素。考虑到成本、灵活性和兼容性，我们预计PACIFIC可以为任何使用传统免疫荧光平台的人提供深入的表型能力。

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来源期刊

ACS Bio & Med Chem Au 药物、生物、化学-

CiteScore

4.10

自引率

0.00%

发文量

期刊介绍： ACS Bio & Med Chem Au is a broad scope open access journal which publishes short letters comprehensive articles reviews and perspectives in all aspects of biological and medicinal chemistry. Studies providing fundamental insights or describing novel syntheses as well as clinical or other applications-based work are welcomed.This broad scope includes experimental and theoretical studies on the chemical physical mechanistic and/or structural basis of biological or cell function in all domains of life. It encompasses the fields of chemical biology synthetic biology disease biology cell biology agriculture and food natural products research nucleic acid biology neuroscience structural biology and biophysics.The journal publishes studies that pertain to a broad range of medicinal chemistry including compound design and optimization biological evaluation molecular mechanistic understanding of drug delivery and drug delivery systems imaging agents and pharmacology and translational science of both small and large bioactive molecules. Novel computational cheminformatics and structural studies for the identification (or structure-activity relationship analysis) of bioactive molecules ligands and their targets are also welcome. The journal will consider computational studies applying established computational methods but only in combination with novel and original experimental data (e.g. in cases where new compounds have been designed and tested).Also included in the scope of the journal are articles relating to infectious diseases research on pathogens host-pathogen interactions therapeutics diagnostics vaccines drug-delivery systems and other biomedical technology development pertaining to infectious diseases.