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Nat. Commun., Published 05 May 2022 | https://doi.org/10.1038/s41467-022-30168-z PICASSO allows ultra-multiplexed fluorescence imaging of spatially overlapping proteins without reference spectra measurements

Junyoung Seo^1,4, Yeonbo Sim^2,4, Jeewon Kim^3,4, Hyunwoo Kim¹, In Cho¹, Hoyeon Nam¹, Young-Gyu Yoon^3,* & Jae-Byum Chang^1,*

¹Department of Materials Science and Engineering, Korea Advanced Institute of Science and Technology, Daejeon, Korea. ²Department of Biomedical Engineering, Sungkyunkwan University, Suwon, Korea. ³School of Electrical Engineering, Korea Advanced Institute of Science and Technology, Daejeon, Korea. ⁴These authors contributed equally: Junyoung Seo, Yeonbo Sim, Jeewon Kim.

^*Corresponding author.

Abstract

Ultra-multiplexed fluorescence imaging requires the use of spectrally overlapping fluorophores to label proteins and then to unmix the images of the fluorophores. However, doing this remains a challenge, especially in highly heterogeneous specimens, such as the brain, owing to the high degree of variation in the emission spectra of fluorophores in such specimens. Here, we propose PICASSO, which enables more than 15-color imaging of spatially overlapping proteins in a single imaging round without using any reference emission spectra. PICASSO requires an equal number of images and fluorophores, which enables such advanced multiplexed imaging, even with bandpass filter-based microscopy. We show that PICASSO can be used to achieve strong multiplexing capability in diverse applications. By combining PICASSO with cyclic immunofluorescence staining, we achieve 45-color imaging of the mouse brain in three cycles. PICASSO provides a tool for multiplexed imaging with high accessibility and accuracy for a broad range of researchers.

논문정보

형식Research article
게재일2022년 05월 (BRIC 등록일 2022-06-08)
연구진국내 연구진
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