Optical detection and spectroscopy of single molecules and single nanoparticles have been achieved at room temperature with the use of surface-enhanced Raman scattering. Individual silver colloidal nanoparticles were screened from a large heterogeneous population for special size-dependent properties and were then used to amplify the spectroscopic signatures of adsorbed molecules. For single rhodamine 6G molecules adsorbed on the selected nanoparticles, the intrinsic Raman enhancement factors were on the order of 10 14 to 10 15 , much larger than the ensemble-averaged values derived from conventional measurements. This enormous enhancement leads to vibrational Raman signals that are more intense and more stable than single-molecule fluorescence.

http://science.sciencemag.org/content/sci/275/5303/1102.full.pdf

Probing Single Molecules and Single Nanoparticles by Surface-Enhanced Raman Scattering

Signaling Recognition Events with Fluorescent Sensors and Switches.

Soluble oligomers are common to most amyloids and may represent the primary toxic species of amyloids, like the Aβ peptide in Alzheimer's disease (AD). Here we show that all of the soluble oligomers tested display a common conformation-dependent structure that is unique to soluble oligomers regardless of sequence. The in vitro toxicity of soluble oligomers is inhibited by oligomer-specific antibody. Soluble oligomers have a unique distribution in human AD brain that is distinct from fibrillar amyloid. These results indicate that different types of soluble amyloid oligomers have a common structure and suggest they share a common mechanism of toxicity.

http://science.sciencemag.org/content/sci/300/5618/486.full.pdf

Common Structure of Soluble Amyloid Oligomers Implies Common Mechanism of Pathogenesis

Microfluidic devices are finding increasing application as analytical systems, biomedical devices, tools for chemistry and biochemistry, and systems for fundamental research. Conventional methods of fabricating microfluidic devices have centered on etching in glass and silicon. Fabrication of microfluidic devices in poly(dimethylsiloxane) (PDMS) by soft lithography provides faster, less expensive routes than these conventional methods to devices that handle aqueous solutions. These soft-lithographic methods are based on rapid prototyping and replica molding and are more accessible to chemists and biologists working under benchtop conditions than are the microelectronics-derived methods because, in soft lithography, devices do not need to be fabricated in a cleanroom. This paper describes devices fabricated in PDMS for separations, patterning of biological and nonbiological material, and components for integrated systems.

Fabrication of microfluidic systems in poly(dimethylsiloxane)

The ability to control the size, shape, and material of a surface has reinvigorated the field of surface-enhanced Raman spectroscopy (SERS). Because excitation of the localized surface plasmon resonance of a nanostructured surface or nanoparticle lies at the heart of SERS, the ability to reliably control the surface characteristics has taken SERS from an interesting surface phenomenon to a rapidly developing analytical tool. This article first explains many fundamental features of SERS and then describes the use of nanosphere lithography for the fabrication of highly reproducible and robust SERS substrates. In particular, we review metal film over nanosphere surfaces as excellent candidates for several experiments that were once impossible with more primitive SERS substrates (e.g., metal island films). The article also describes progress in applying SERS to the detection of chemical warfare agents and several biological molecules.

Surface-Enhanced Raman Spectroscopy

A method is described that provides for detection and identification of single molecules in solution. The method is based on fluorescence correlation spectroscopy, which records spatio-temporal correlations among fluctuating light signals, coupled with devices for trapping single molecules in an electric field. This technique is applied to studies of molecular evolution, where it allows fast screening of large mutant spectra in which targets are labeled by specific fluorescent ligands. The method expands the horizon in molecular diagnostics by making it possible to monitor concentrations down to (less than) 10(-15) M without any need for amplification.

https://www.pnas.org/content/pnas/91/13/5740.full.pdf

Sorting single molecules: application to diagnostics and evolutionary biotechnology

An epi-illuminated microscope configuration for use in fluorescence correlation spectroscopy in bulk solutions has been analyzed. For determining the effective sample dimensions the spatial distribution of the molecule detection efficiency has been computed and conditions for achieving quasi-cylindrical sample shape have been derived. Model experiments on translational diffusion of rhodamine 6G have been carried out using strong focusing of the laser beam, small pinhole size and an avalanche photodiode in single photon counting mode as the detector. A considerable decrease in background light intensity and measurement time has been observed. The background light is 40 times weaker than the fluorescence signal from one molecule of Rh6G, and the correlation function with signal-to-noise ratio of 150 can be collected in 1 second. The effect of the shape of the sample volume on the autocorrelation function has been discussed.

Fluorescence correlation spectroscopy with high count rate and low background: analysis of translational diffusion

Fluorescence correlation spectroscopy

http://www.biophysik.physik.uni-muenchen.de/download/biopheinf/seminar/Schwille_1997_JACS.pdf

Rudolf Rigler

Papers

Sorting single molecules: application to diagnostics and evolutionary biotechnology

Fluorescence correlation spectroscopy with high count rate and low background: analysis of translational diffusion

Fluorescence correlation spectroscopy

Dual-color fluorescence cross-correlation spectroscopy for multicomponent diffusional analysis in solution

Fluorescence correlation spectroscopy of triplet states in solution: a theoretical and experimental study