title: Single cell manipulation, analytics, and label-free protein detection in microfluidic devices for systems nanobiology.
abstract: Single cell analytics for proteomic analysis is considered a key method in the framework of systems nanobiology which allows a novel proteomics without being subjected to ensemble-averaging, cell-cycle, or cell-population effects. We are currently developing a single cell analytical method for protein fingerprinting combining a structured microfluidic device with latest optical laser technology for single cell manipulation (trapping and steering), free-solution electrophoretical protein separation, and (label-free) protein detection. In this paper we report on first results of this novel analytical device focusing on three main issues. First, single biological cells were trapped, injected, steered, and deposited by means of optical tweezers in a poly(dimethylsiloxane) microfluidic device and consecutively lysed with SDS at a predefined position. Second, separation and detection of fluorescent dyes, amino acids, and proteins were achieved with LIF detection in the visible (VIS) (488 nm) as well as in the deep UV (266 nm) spectral range for label-free, native protein detection. Minute concentrations of 100 fM injected fluorescein could be detected in the VIS and a first protein separation and label-free detection could be achieved in the UV spectral range. Third, first analytical experiments with single Sf9 insect cells (Spodoptera frugiperda) in a tailored microfluidic device exhibiting distinct electropherograms of a green fluorescent protein-construct proved the validity of the concept. Thus, the presented microfluidic concept allows novel and fascinating single cell experiments for systems nanobiology in the future.
authors.affiliation: Experimental Biophysics and Applied Nanosciences, Physics Department, Bielefeld University, Bielefeld, Germany.
authors.forename: Wibke
authors.initials: W
authors.lastname: Hellmich
authors.forename: Christoph
authors.initials: C
authors.lastname: Pelargus
authors.forename: Kai
authors.initials: K
authors.lastname: Leffhalm
authors.forename: Alexandra
authors.initials: A
authors.lastname: Ros
authors.forename: Dario
authors.initials: D
authors.lastname: Anselmetti
mesh_terms: D000818:Animals; D002460:Cell Line; D002477:Cells / Q000737:chemistry; D004586:Electrophoresis; D005456:Fluorescent Dyes / Q000032:analysis; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D036103:Nanotechnology / Q000295:instrumentation*; D011506:Proteins / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D018411:Spodoptera / Q000166:cytology; D014466:Ultraviolet Rays
chemical_list: D005456:Fluorescent Dyes; D011506:Proteins