Heme proteins, such as hemoglobins and cytochromes, play important roles in various biological processes. Here we employ the two-photon excited photothermal effect as a contrast mechanism to map heme proteins distribution. Particularly, both a thermal lens scheme and a high-frequency modulation are utilized to enhance the signal-to-noise ratio. We demonstrate label-free imaging of individual red blood cells, subcellular distribution of cytochromes in live mammalian cells, and the microvascular networks in mouse ear tissue and in a zebrafish gill.

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Direct cellular imaging of the localization and dynamics of biomolecules helps to understand their function and reveals novel mechanisms at the single-cell resolution. In contrast to routine fluorescent-protein-based protein imaging, technology for RNA imaging remains less well explored because of the lack of enabling technology. Herein, we report the development of an aptamer-initiated fluorescence complementation (AiFC) method for RNA imaging by engineering a green fluorescence protein (GFP)-mimicking turn-on RNA aptamer, Broccoli, into two split fragments that could tandemly bind to target mRNA. When genetically encoded in cells, endogenous mRNA molecules recruited Split-Broccoli and brought the two fragments into spatial proximity, which formed a fluorophore-binding site in situ and turned on fluorescence. Significantly, we demonstrated the use of AiFC for high-contrast and real-time imaging of endogenous RNA molecules in living mammalian cells. We envision wide application and practical utility of this enabling technology to in vivo single-cell visualization and mechanistic analysis of macromolecular interactions.

Cell-membrane-camouflaged nanoparticles (CMC-NPs) have been increasingly exploited to develop various therapeutic tools due to their high biocompatibility and cell-type-specific tumor-targeting properties. However, the molecular mechanism of CMC-NPs for homotypic targeting remains elusive. Here, we develop a plasmonic imaging method by coating gold nanoparticles (AuNPs) with cancer cell membranes and perform plasmonic imaging of the interactions between CMC-NPs and living cells at the single-cell level. Quantitative analysis of CMC-NPs in a different clustering status reveals that the presence of cell membranes on CMC-NPs results in a 7-fold increase in homotypic cell delivery and nearly 2 orders of magnitude acceleration of the intracellular agglomeration process. Significantly, we identify that integrin αvβ3, a cell surface receptor abundantly expressed in tumor cells, is critical for the selective cell recognition of CMC-NPs. We thus establish a single-cell plasmonic imaging platform for probing NP–cell interactions, which sheds new light on the therapeutic applications of CMC-NPs.