title: Nine orders of magnitude dynamic range: picomolar to millimolar concentration measurement in capillary electrophoresis with laser induced fluorescence detection employing cascaded avalanche photodiode photon counters.
abstract: The dynamic range of capillary electrophoresis analysis is ultimately limited by molecular shot noise at low concentrations and by concentration-induced band broadening at high concentrations. We report a system that approaches these fundamental limits. A laser-induced fluorescence detector is reported that employs a cascade of four fiber-optic beam splitters connected in series to generate a primary signal and four attenuated signals, each monitored by a single-photon counting avalanche photodiode. Appropriate scaling of the signals from the five photodiodes produces a linear optical calibration curve for 5-carboxyl-tetramethylrhodamine from the concentration detection limit of 1 pM to the upper limit of 1 mM. Mass detection limits are 120 yoctomoles (70 molecules) injected into the instrument. The very-wide dynamic range instrument was used to study the metabolic products of the fluorescently labeled glycosphingolipid tetramethylrhodamine labeled GM1 (GM1-TMR) produced by single cells isolated from the rat cerebellum.
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, Indiana 46556, USA.
authors.forename: Oluwatosin O
authors.initials: OO
authors.lastname: Dada
authors.forename: David C
authors.initials: DC
authors.lastname: Essaka
authors.forename: Ole
authors.initials: O
authors.lastname: Hindsgaul
authors.forename: Monica M
authors.initials: MM
authors.lastname: Palcic
authors.forename: Jillian
authors.initials: J
authors.lastname: Prendergast
authors.forename: Ronald L
authors.initials: RL
authors.lastname: Schnaar
authors.forename: Norman J
authors.initials: NJ
authors.lastname: Dovichi
mesh_terms: D000818:Animals; D002138:Calibration; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism; D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D006017:Glycolipids / Q000302:isolation & purification / Q000378:metabolism; D007834:Lasers; D057230:Limit of Detection; D017785:Photons; D051381:Rats; D059010:Single-Cell Analysis; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D006017:Glycolipids