TY  - JOUR
AU  - Dertinger, T.
AU  - Pacheco, V.
AU  - von der Hocht, I.
AU  - Hartmann, R.
AU  - Gregor, I.
AU  - Enderlein, J.
TI  - Two focus fluorescence correlation spectroscopy: A new tool for accurate and absolute diffusion measurements
JO  - ChemPhysChem
VL  - 8
SN  - 1439-4235
CY  - Weinheim
PB  - Wiley-VCH Verl.
M1  - PreJuSER-58183
SP  - 433 - 443
PY  - 2007
N1  - Record converted from VDB: 12.11.2012
AB  - We present a new method to measure absolute diffusion coefficients at nanomolar concentrations with high precision. Based on a modified fluorescence correlation spectroscopy (FCS)-setup, this method is improved by introducing an external ruler for measuring the diffusion time by generating two laterally shifted and overlapping laser foci at a fixed and known distance. Data fitting is facilitated by a new two-parameter model to describe the molecule detection function (MDF). We present a recorded MDF and show the excellent agreement with the fitting model. We measure the diffusion coefficient of the red fluorescent dye Atto655 under various conditions and compare these values with a value achieved by gradient pulsed field NMR (GPF NMR). From these measurements we conclude, that the new measurement scheme is robust against optical and photophysical artefacts which are inherent to standard FCS. With two-focus-FCS, the diffusion coefficient of 4.26 x 10(-6) cm2s(-1) for Atto655 in water at 25 degrees C compares well with the GPF NMR value of 4.28 x 10(-6) cm2s(-1).
KW  - Diffusion
KW  - Sensitivity and Specificity
KW  - Solutions
KW  - Spectrometry, Fluorescence: methods
KW  - Viscosity
KW  - Solutions (NLM Chemicals)
KW  - J (WoSType)
LB  - PUB:(DE-HGF)16
C6  - pmid:17269116
UR  - <Go to ISI:>//WOS:000244496200011
DO  - DOI:10.1002/cphc.200600638
UR  - https://juser.fz-juelich.de/record/58183
ER  -