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SHR Neuro Cancer Cardio Lipid Metab Microb

Földes-Papp, Z; Liao, SC; You, T; Terpetschnig, E; Barbieri, B.
Confocal fluctuation spectroscopy and imaging.
Curr Pharm Biotechnol. 2010; 11(6):639-653 Doi: 10.2174/138920110792246618
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Leading authors Med Uni Graz

Földes-Papp Zeno

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Abstract:

Currently, work with subnanomolar concentrations is routine while femtomolar and even single-molecule studies are possible with some efforts getting high on single-molecule biophysics and biochemistry. Methodological breakthroughs, such as reducing the background light contribution in single-molecule studies, which has plagued many studies of molecular fluorescence in dilute solution, and particularly in live cells, have recently described by us. We first demonstrated how optimized time-gating of the fluorescence signal, together with time-correlated, single-photon counting, can be used to substantially boost the experimental signal-to-noise ratio about 140-fold, making it possible to measure analyte concentrations that are as low as 15 pM. By detection of femtomolar bulk concentrations, confocal microsopy has the potential to address the observation of one and the same molecule in dilute solution without immobilization or hydrodynamic/electrokinetic focusing at longer observation times than currently available. We present relevant physics. The equations are derived using Einstein's approach showing how it fits with Fick's law and the autocorrelation function. An improved technology is being developed at ISS for femtomolar microscopy. The general concepts and provided experimental examples should help to compare our approach to those used in conventional confocal microscopy.

Find related publications in this database (using NLM MeSH Indexing)

Animals -

Computer Simulation -

Humans -

Image Interpretation, Computer-Assisted - methods

Microscopy, Fluorescence - methods

Models, Biological -

Models, Chemical -

Molecular Imaging - methods

Find related publications in this database (Keywords)

Fluctuation microscopy

fluctuation spectroscopy

fluctuation imaging

lifetime imaging

FLIM

fast FLIM

single-molecule detection in dilute solution

live cell

physical concepts

Brownian motion

optimized ISS fluorescence fluctuation system ALBA-JZ (TM)

how-to-do approach

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