A single fluorescent probe enables clearly discriminating and simultaneously imaging liquid-ordered and liquid-disordered microdomains in plasma membrane of living cells.

Liquid-ordered (Lo) and liquid-disordered (Ld) microdomains in plasma membrane play different yet essential roles in various bioactivities. However, discrimination of the two microdomains in living cells is difficult, due to the similarity in their constituents and structures. Up to now, polarity sensitive probes are the only tool for imaging the two microdomains, but their small difference between emission spectra in the two microdomains (less than 50 nm) limited their application in living cells. In this work, we first presented an aggregation/monomer type of fluorescent probe (2,7-9E-BHVC12) with much larger separation in emission wavelength (up to 100 nm), for dual-color visualizing the two membrane microdomains in living cells. The probe can form red-emissive aggregates and yellow-emissive monomers when induced by Lo and Ld microdomains, respectively, and thus enables clear visualization of the two membrane microdomains in living cells with dual colors, and thus high-fidelity images of substructures of plasma membrane have been obtained. According to the images of three kinds of normal cells and three kinds of cancer cells stained with 2,7-9E-BHVC12, significant difference in plasma membrane microstructure of cancer cells was found. In terms of 2,7-9E-BHVC12, normal cells were mainly consisted of either Lo or Ld microdomains all over their membranes, while cancer cells all clearly display coexistence of Lo and Ld membrane microdomains. Therefore, 2,7-9E-BHVC12 can serve as a powerful tool for studies of membrane microdomains, and the different results of normal and cancer cells would also deepen our understanding in cancer science.