Rapid Multiplexed Imaging of Cell-Surface Cancer Biomarkers in Fresh Tissues with Targeted SERS Nanoparticles

Our lab is developing miniature Raman imaging systems and topical-staining protocols to rapidly image cell-surface biomarkers in fresh tissues. In particular, this work employs targeted surface-enhanced Raman scattering (SERS) nanoparticles (NPs) to enable the sensitive and multiplexed detection of a large number of cell-surface biomarkers of cancer. The SERS NPs were functionalized with different targeting antibodies, and their biomarker detection capability was investigated via in vitro and ex vivo experiments with cells and tissues. Here, we design SERS NPs to specifically target the cancer biomarker EGFR upon topical application on cells and tissues. In vitro flow cytometry with fluorescent SERS NPs reveals a high ratio of specific versus nonspecific binding for the tumor cell lines A431 (skin cancer), U251 (glioma) and SkBr3 (breast cancer). For tissue imaging, we have developed a fiber-optic-based spectral detection probe, with 785-nm laser illumination, for rapid detection of SERS NPs with sub-millimeter spatial resolution. Based on the spectral detection probe, multiple imaging systems were customized for rapid tissue phenotyping such as a comprehensive rotational scanning endoscope for in vivo imaging of the rat esophagus and a raster-scanning device for intraoperative imaging of breast tissue margins. Ex vivo experiments were performed to develop a strategy for the rapid detection of multiple cell-surface biomarkers following a brief (5-10 min) topical application of SERS NPs on tissues. By developing high-affinity targeted SERS NPs, sensitive spectral-imaging devices, and an optimized topical-delivery protocol, we demonstrate a ratio metric method to rapidly quantify the specific binding of biomarker-targeted NPs on fresh tissues, thereby eliminating the ambiguities that often arise due to nonspecific sources of contrast. These tools will enable multiplexed molecular imaging for the early detection of epithelial cancers, rapid surgical guidance, and moni- oring the molecular response to treatments.