title: Optical Clearing Delivers Ultrasensitive Hyperspectral Dark-Field Imaging for Single-Cell Evaluation.
abstract: A single-cell optical clearing methodology is developed and demonstrated in hyperspectral dark-field microscopy (HSDFM) and imaging of plasmonic nanoprobes. Our strategy relies on a combination of delipidation and refractive index (RI) matching with highly biocompatible and affordable agents. Before applying the RI-matching solution, the delipidation step by using a mild solvent effectively eliminates those high-density, lipid-enriched granular structures which emit strong scattering. Upon treatment, the background scattering from cellular organelles could be repressed to a negligible level while the scattering signals from plasmonic nanomaterials increase, leading to a significant improvement of the signal-to-noise ratio (SNR). With this method established, the versatility and applicability of HSDFM are greatly enhanced. In our demonstration, quantitative mapping of the dimerization-activated receptor kinase HER2 is achieved in a single cancer cell by a nonfluorescent approach. High-resolution imaging for oncogenic mRNAs, namely ER, PR, and HER2, is performed with single labeling. More importantly, in situ multiplex detection of mRNA and protein is made possible by HSDFM since it overcomes the difficulties of complex staining and signal imbalance suffered by the conventional optical imaging. Last, we show that with optical clearing, characterization of intracellularly grown gold particulates is accomplished at an unprecedented spatiotemporal resolution. Taken together, the uniqueness of optical clearing and HSDFM is expected to open ample avenues for single-cell studies and biomedical engineering.
authors.affiliation: Department of Agricultural and Biological Engineering, Bindley Bioscience Center and Birck Nanotechnology Center, Purdue Center for Cancer Research, Purdue University , West Lafayette, Indiana 47907, United States.
authors.forename: Yi
authors.initials: Y
authors.lastname: Cui
authors.affiliation: Department of Agricultural and Biological Engineering, Bindley Bioscience Center and Birck Nanotechnology Center, Purdue Center for Cancer Research, Purdue University , West Lafayette, Indiana 47907, United States.
authors.forename: Xiaolei
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Agricultural and Biological Engineering, Bindley Bioscience Center and Birck Nanotechnology Center, Purdue Center for Cancer Research, Purdue University , West Lafayette, Indiana 47907, United States.
authors.forename: Wen
authors.initials: W
authors.lastname: Ren
authors.affiliation: Department of Agricultural and Biological Engineering, Bindley Bioscience Center and Birck Nanotechnology Center, Purdue Center for Cancer Research, Purdue University , West Lafayette, Indiana 47907, United States.
authors.forename: Jing
authors.initials: J
authors.lastname: Liu
authors.affiliation: Department of Agricultural and Biological Engineering, Bindley Bioscience Center and Birck Nanotechnology Center, Purdue Center for Cancer Research, Purdue University , West Lafayette, Indiana 47907, United States.
authors.forename: Joseph
authors.initials: J
authors.lastname: Irudayaraj
mesh_terms: D045744:Cell Line, Tumor; D006801:Humans; D008027:Light; D008853:Microscopy / Q000379:methods; D049329:Nanostructures / Q000737:chemistry; D009369:Neoplasms / Q000000981:diagnostic imaging; D061848:Optical Imaging / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000032:analysis; D012031:Refractometry; D012542:Scattering, Radiation; D059629:Signal-To-Noise Ratio; D059010:Single-Cell Analysis
chemical_list: D012333:RNA, Messenger; D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human
keywords: hyperspectral dark-field microscopy; optical clearing; plasmonic nanomaterials; single-cell analysis; single-molecule detection