Intracellular organelles and components are highly heterogeneous at the nanometer-scale in terms of both physical structure and chemical composition. The structural heterogeneity can be readily visualized by super-resolution microscopy methods such as Stochastic Optical Reconstruction Microscopy (STORM). However, investigating the nanoscale heterogeneity in the chemical composition remains a challenge.<br/>Here we combined spectrally resolved STORM (SR-STORM), a new method we recently developed, with fluorophores that are spectrally responsive to local polarity, to investigate the nanoscale heterogeneity in cellular membranes. The spectrum and position of every single fluorophore molecule in a labeled live cell were simultaneously recorded through SR-STORM. Combined with the membrane polarity marker, SR-STORM enables us to resolve the spectral difference between plasma membrane and the intracellular membranes of mitochondria and endoplasmic reticulum (ER), along with their nanoscale structures. Single-molecule spectra on different types of membranes were successfully distinguished based on their emission spectra. Additionally, spectral shift of the single-molecule spectra was observed upon drug treatments. Therefore, this approach provides a powerful way to investigate the ultrafine structures and local chemical composition of cellular membranes.

Probing intracellular micro-environments through spectrally resolved STORM

253rd National Meeting (2017)

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