Multiplex Protein Imaging Through PACIFIC: Photoactive Immunofluorescence with Iterative Cleavage

Overview

Journal ACS Bio Med Chem Au

Publisher American Chemical Society

Specialty Biochemistry

Date 2023 Jun 26

PMID 37363079

Authors

Fei Ji

Moises Hur

Sungwon Hur

Siwen Wang

Priyanka Sarkar

Shiqun Shao

Desiree Aispuro

Xu Cong

Yanhao Hu

Zhonghan Li

Min Xue

Affiliations

Soon will be listed here.

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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