2014, Neugebauer, U., Kurz, C., Bocklitz, T., Berger, T., Velten, T., Clement, J.H., Krafft, C., Popp, J.

Circulating tumor cells (CTCs) from blood of cancer patients are valuable prognostic markers and enable monitoring responses to therapy. The extremely low number of CTCs makes their isolation and characterization a major technological challenge. For label-free cell identification a novel combination of Raman spectroscopy with a microhole array platform is described that is expected to support high-throughput and multiplex analyses. Raman spectra were registered from regularly arranged cells on the chip with low background noise from the silicon nitride chip membrane. A classification model was trained to distinguish leukocytes from myeloblasts (OCI-AML3) and breast cancer cells (MCF-7 and BT-20). The model was validated by Raman spectra of a mixed cell population. The high spectral quality, low destructivity and high classification accuracy suggests that this approach is promising for Raman activated cell sorting.

2015, Peckys, Diana B., Korf, Ulrike, de Jonge, Niels

The formation of HER2 homodimers plays an important role in breast cancer aggressiveness and progression; however, little is known about its localization. We have studied the intra- and intercellular variation of HER2 at the single-molecule level in intact SKBR3 breast cancer cells. Whole cells were visualized in hydrated state with correlative fluorescence microscopy and environmental scanning electron microscopy (ESEM). The locations of individual HER2 receptors were detected using an anti-HER2 affibody in combination with a quantum dot (QD), a fluorescent nanoparticle. Fluorescence microscopy revealed considerable differences of HER2 membrane expression between individual cells and between different membrane regions of the same cell (that is, membrane ruffles and flat areas). Subsequent ESEM of the corresponding cellular regions provided images of individually labeled HER2 receptors. The high spatial resolution of 3 nm and the close proximity between the QD and the receptor allowed quantifying the stoichiometry of HER2 complexes, distinguishing between monomers, dimers, and higher-order clusters. Downstream data analysis based on calculating the pair correlation function from receptor positions showed that cellular regions exhibiting membrane ruffles contained a substantial fraction of HER2 in homodimeric state. Larger-order clusters were also present. Membrane areas with homogeneous membrane topography, on the contrary, displayed HER2 in random distribution. Second, HER2 homodimers appeared to be absent from a small subpopulation of cells exhibiting a flat membrane topography, possibly resting cells. Local differences in homodimer presence may point toward functional differences with possible relevance for studying metastasis and drug response.