The advent of image-activated cell sorting and imaging-based cell picking has advanced our knowledge and exploitation of biological systems in the last decade. Unfortunately, they generally rely on fluorescent labeling for cellular phenotyping, an indirect measure of the molecular landscape in the cell, which has critical limitations. Here we demonstrate Raman image-activated cell sorting by directly probing chemically specific intracellular molecular vibrations via ultrafast multicolor stimulated Raman scattering (SRS) microscopy for cellular phenotyping. Specifically, the technology enables real-time SRS-image-based sorting of single live cells with a throughput of up to~100 events per second without the need for fluorescent labeling. To show the broad utility of the technology, we show its applicability to diverse cell types and sizes. The technology is highly versatile and holds promise for numerous applications that are previously difficult or undesirable with fluorescence-based technologies.
The holotype (MHM-K2) of the Eocene cheloniine Tasbacka danica is arguably one of the best preserved juvenile fossil sea turtles on record. Notwithstanding compactional flattening, the specimen is virtually intact, comprising a fully articulated skeleton exposed in dorsal view. MHM-K2 also preserves, with great fidelity, soft tissue traces visible as a sharply delineated carbon film around the bones and marginal scutes along the edge of the carapace. Here we show that the extraordinary preservation of the type of T. danica goes beyond gross morphology to include ultrastructural details and labile molecular components of the once-living animal. Haemoglobin-derived compounds, eumelanic pigments and proteinaceous materials retaining the immunological characteristics of sauropsid-specific β-keratin and tropomyosin were detected in tissues containing remnant melanosomes and decayed keratin plates. The preserved organics represent condensed remains of the cornified epidermis and, likely also, deeper anatomical features, and provide direct chemical evidence that adaptive melanism – a biological means used by extant sea turtle hatchlings to elevate metabolic and growth rates – had evolved 54 million years ago.
By virtue of the combined merits of flow cytometry and fluorescence microscopy, imaging flow cytometry (IFC) has become an established tool for cell analysis in diverse biomedical fields such as cancer biology, microbiology, immunology, hematology, and stem cell biology. However, the performance and utility of IFC are severely limited by the fundamental trade-off between throughput, sensitivity, and spatial resolution. Here we present an optomechanical imaging method that overcomes the trade-off by virtually freezing the motion of flowing cells on the image sensor to effectively achieve 1000 times longer exposure time for microscopygrade fluorescence image acquisition. Consequently, it enables high-throughput IFC of single cells at >10,000 cells s −1 without sacrificing sensitivity and spatial resolution. The availability of numerous information-rich fluorescence cell images allows high-dimensional statistical analysis and accurate classification with deep learning, as evidenced by our demonstration of unique applications in hematology and microbiology.
Purpose: HER3 is a compelling target for cancer treatment; however, no HER3-targeted therapy is currently clinically available. Here, we produced U3-1402, an anti-HER3 antibody-drug conjugate with a topoisomerase I inhibitor exatecan derivative (DXd), and systematically investigated its targeted drug delivery potential and antitumor activity in preclinical models.Experimental Design: In vitro pharmacologic activities and the mechanisms of action of U3-1402 were assessed in several human cancer cell lines. Antitumor activity of U3-1402 was evaluated in xenograft mouse models, including patientderived xenograft (PDX) models. Safety assessments were also conducted in rats and monkeys.Results: U3-1402 showed HER3-specific binding followed by highly efficient cancer cell internalization. Subsequently, U3-1402 was translocated to the lysosome and released its payload DXd. While U3-1402 was able to inhibit HER3activated signaling similar to its naked antibody patritumab, the cytotoxic activity of U3-1402 in HER3-expressing cells was predominantly mediated by released DXd through DNA damage and apoptosis induction. In xenograft mouse models, U3-1402 exhibited dose-dependent and HER3-dependent antitumor activity. Furthermore, U3-1402 exerted potent antitumor activity against PDX tumors with HER3 expression. Acceptable toxicity was noted in both rats and monkeys.Conclusions: U3-1402 demonstrated promising antitumor activity against HER3-expressing tumors with tolerable safety profiles. The activity of U3-1402 was driven by HER3mediated payload delivery via high internalization into tumor cells.
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