Sequential, Multiplexed Immunofluorescent Imaging of Live Cells Based on DNA-Mediated Reversible Fluorophore Attachment/Detachment with Antibodies.

ABSTRACT: Given the spectral overlap of fluorophores, traditional immunofluorescence imaging is limited by the number of proteins that can be imaged simultaneously. Although sequential imaging techniques have been proposed, in which repeated staining and destaining are performed to obtain the merged image of several proteins, they are applied only to fixed cells presumably due to their harsh conditions. Therefore, observation and analysis of live cells have not been achieved with the sequential imaging approach. In this study, we develop a sequential, multiplexed immunofluorescence imaging method for live cells using DNA as a detachable linker to bind antibodies to fluorophores. The use of toehold-mediated strand displacement of DNAs enables the attachment and detachment of fluorophores under mild p

SUBMITTER: Xu L 

PROVIDER: S-EPMC12457991 | biostudies-literature | 2025 Sep

REPOSITORIES: biostudies-literature