title: Six-color time-resolved Förster resonance energy transfer for ultrasensitive multiplexed biosensing.
abstract: Simultaneous monitoring of multiple molecular interactions and multiplexed detection of several diagnostic biomarkers at very low concentrations have become important issues in advanced biological and chemical sensing. Here we present an optically multiplexed six-color Förster resonance energy transfer (FRET) biosensor for simultaneous monitoring of five different individual binding events. We combined simultaneous FRET from one Tb complex to five different organic dyes measured in a filter-based time-resolved detection format with a sophisticated spectral crosstalk correction, which results in very efficient background suppression. The advantages and robustness of the multiplexed FRET sensor were exemplified by analyzing a 15-component lung cancer immunoassay involving 10 different antibodies and five different tumor markers in a single 50 μL human serum sample. The multiplexed biosensor offers clinically relevant detection limits in the low picomolar (ng/mL) concentration range for all five markers, thus providing an effective early screening tool for lung cancer with the possibility of distinguishing small-cell from non-small-cell lung carcinoma. This novel technology will open new doors for multiple biomarker diagnostics as well as multiplexed real-time imaging and spectroscopy.
authors.affiliation: Physical Chemistry, University of Potsdam, 14476 Potsdam-Golm, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Geißler
authors.forename: Stefan
authors.initials: S
authors.lastname: Stufler
authors.forename: Hans-Gerd
authors.initials: HG
authors.lastname: Löhmannsröben
authors.forename: Niko
authors.initials: N
authors.lastname: Hildebrandt
mesh_terms: D014408:Biomarkers, Tumor / Q000097:blood*; D015374:Biosensing Techniques; D002289:Carcinoma, Non-Small-Cell Lung / Q000175:diagnosis*; D003116:Color; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D008175:Lung Neoplasms / Q000175:diagnosis*; D055752:Small Cell Lung Carcinoma / Q000175:diagnosis*; D013725:Terbium / Q000737:chemistry; D013997:Time Factors
chemical_list: D014408:Biomarkers, Tumor; D005456:Fluorescent Dyes; D013725:Terbium