Rxivist logo

Rxivist combines preprints from bioRxiv with data from Twitter to help you find the papers being discussed in your field. Currently indexing 52,519 bioRxiv papers from 243,473 authors.

Multiplexed confocal and super-resolution fluorescence imaging of cytoskeletal and neuronal synapse proteins

By Syuan-Ming Guo, Remi Veneziano, Simon Gordonov, Li Li, Demian Park, Anthony B Kulesa, Paul C Blainey, Jeffrey R Cottrell, Edward S Boyden, Mark Bathe

Posted 25 Feb 2017
bioRxiv DOI: 10.1101/111625

Neuronal synapses contain dozens of protein species whose expression levels and localizations are key determinants of synaptic transmission and plasticity. The spectral properties of fluorophores used in conventional microscopy limit the number of measured proteins to four species within a given sample. The ability to perform high-throughput confocal or super-resolution imaging of many proteins simultaneously without limitation in target number imposed by this spectral limit would enable large-scale characterization of synaptic protein networks in situ. Here, we introduce PRISM: Probe-based Imaging for Sequential Multiplexing, a method that sequentially utilizes either high affinity Locked Nucleic Acid (LNA) or low affinity DNA probes to enable diffraction-limited confocal and PAINT-based super-resolution imaging. High-affinity LNA probes offer high-throughput, confocal-based imaging compared with PAINT, which uses low affinity probes to realize localization-based super-resolution imaging. Simultaneous immunostaining of all targets is performed prior to imaging, followed by sequential LNA/DNA probe exchange that requires only minutes under mild wash conditions. We apply PRISM to quantify the co-expression levels and nanometer-scale organization of one dozen cytoskeletal and synaptic proteins within individual neuronal synapses. Our approach is scalable to dozens of target proteins and is compatible with high-content screening platforms commonly used to interrogate phenotypic changes associated with genetic and drug perturbations in a variety of cell types.

Download data

  • Downloaded 3,924 times
  • Download rankings, all-time:
    • Site-wide: 536 out of 52,519
    • In bioengineering: 9 out of 976
  • Year to date:
    • Site-wide: 3,893 out of 52,519
  • Since beginning of last month:
    • Site-wide: 4,758 out of 52,519

Altmetric data


Downloads over time

Distribution of downloads per paper, site-wide


Sign up for the Rxivist weekly newsletter! (Click here for more details.)


News