Fluorescence correlation spectroscopy was performed by focusing in
a droplet on a cleaned coverslip on top of a confocal microscope.
The excitation source was 600 nm pulsed light (77 MHz) selected from
a continuum laser (SuperK Extreme EXB-6 with a SuperK SELECT wavelength
selector from NKT Photonics, 290 nW measured on the objective). A
600 nm band-pass filter (Semrock FF01-600/14-25) was used to clean
up excitation light. The laser beam (a round-top-hat-like beam profile
was used) was focused through an Olympus IX71 microscope by the oil
immersion objective (Olympus UPLFLN 100×, 1.3 NA). Scattering
from the excitation source was removed using a dichroic mirror (Semrock
LPD01-633RS) and a long-pass filter (Semrock LP02-633RU-25). The fluorescence
signal was collected through a 100 μm pinhole by an avalanche
photodiode (APD, PerkinElmer CD3226). The signals from the APD were
recorded by an SPC-830 card (Becker & Hickl). Purified DNA-AgNC
solutions were diluted in deionized water to cover a wide range of
concentrations. Atto633 (BioReagent, Sigma-Aldrich) was diluted in
deionized water (concentrations ranging between ∼5.7 and ∼0.63
nM) and used as a reference dye for excitation-volume determination.
All of the FCS measurements were performed at room temperature.
a droplet on a cleaned coverslip on top of a confocal microscope.
The excitation source was 600 nm pulsed light (77 MHz) selected from
a continuum laser (SuperK Extreme EXB-6 with a SuperK SELECT wavelength
selector from NKT Photonics, 290 nW measured on the objective). A
600 nm band-pass filter (Semrock FF01-600/14-25) was used to clean
up excitation light. The laser beam (a round-top-hat-like beam profile
was used) was focused through an Olympus IX71 microscope by the oil
immersion objective (Olympus UPLFLN 100×, 1.3 NA). Scattering
from the excitation source was removed using a dichroic mirror (Semrock
LPD01-633RS) and a long-pass filter (Semrock LP02-633RU-25). The fluorescence
signal was collected through a 100 μm pinhole by an avalanche
photodiode (APD, PerkinElmer CD3226). The signals from the APD were
recorded by an SPC-830 card (Becker & Hickl). Purified DNA-AgNC
solutions were diluted in deionized water to cover a wide range of
concentrations. Atto633 (BioReagent, Sigma-Aldrich) was diluted in
deionized water (concentrations ranging between ∼5.7 and ∼0.63
nM) and used as a reference dye for excitation-volume determination.
All of the FCS measurements were performed at room temperature.