title: Probing Low-Copy-Number Proteins in a Single Living Cell.
abstract: Single-cell analysis techniques are essential for understanding the microheterogeneity and functions of cells. Low-copy-number proteins play important roles in cell functioning, but their measurement in single cells remains challenging. Herein, we report an approach, called plasmonic immunosandwich assay (PISA), for probing low-copy-number proteins in single cells. This approach combined in vivo immunoaffinity extraction and plasmon-enhanced Raman scattering (PERS). Target proteins were specifically extracted from the cells by microprobes modified with monoclonal antibody or molecularly-imprinted polymer (MIP), followed by labeling with Raman-active nanotags. The PERS detection, with Raman intensity enhanced by 9 orders of magnitude, provided ultrasensitive detection at the single-molecule level. Using this approach, we found that alkaline phosphatase and survivin were expressed in distinct levels in cancer and normal cells, and that extended culture passage resulted in reduced expression of survivin. We further developed acupuncture needle-based PISA for probing low-copy-number proteins in living bodies.
authors.affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, 163 Xianlin Avenue, Nanjing, 210023, China.
authors.forename: Jia
authors.initials: J
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, 163 Xianlin Avenue, Nanjing, 210023, China.
authors.forename: Danyang
authors.initials: D
authors.lastname: Yin
authors.affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, 163 Xianlin Avenue, Nanjing, 210023, China.
authors.forename: Shuangshou
authors.initials: S
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, 163 Xianlin Avenue, Nanjing, 210023, China.
authors.forename: Hong-Yuan
authors.initials: HY
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, 163 Xianlin Avenue, Nanjing, 210023, China. zhenliu@nju.edu.cn.
authors.forename: Zhen
authors.identifier: 0000-0002-8440-2554
authors.initials: Z
authors.lastname: Liu
mesh_terms: D015374:Biosensing Techniques; D006801:Humans; D007118:Immunoassay; D054802:Molecular Imprinting; D011108:Polymers / Q000737:chemistry; D011506:Proteins / Q000032:analysis*; D059010:Single-Cell Analysis; D013059:Spectrum Analysis, Raman
chemical_list: D011108:Polymers; D011506:Proteins
keywords: Raman spectroscopy; imprinting; proteins; single-cell analysis; single-molecule detection