Chemical analysis in vivo and in vitro by Raman spectroscopy--from single cells to humans.

The gold standard for clinical diagnostics of tissues is immunofluorescence staining. Toxicity of many fluorescent dyes precludes their application in vivo. Raman spectroscopy, a chemically specific, label-free diagnostic technique, is rapidly gaining acceptance as a powerful alternative.

An adaptable, portable microarray reader for biodetection.

We have developed an inexpensive portable microarray reader that can be applied to standard microscope slide-based arrays and other array formats printed on chemically modified surfaces. Measuring only 19 cm in length, the imaging device is portable and may be applicable to both triage and clinical settings. For multiplexing and adaptability purposes, it can be modified to work with multiple excitation/emission wavelengths.

Nano-biophotonics: new tools for chemical nano-analytics.

The nondestructive chemical analysis of biological processes in the crowded intracellular environment, at cellular membranes, and between cells with a spatial resolution well beyond the diffraction limit is made possible through Nano-Biophotonics. A number of sophisticated schemes employing nanoparticles, nano-apertures, or shaping of the probe volume in the far field have significantly extended our knowledge about lipid rafts, macromolecular complexes, such as chromatin, vesicles, and cellular organelles, and their interactions and trafficking within the cell. Here, I review some of the most recent developments in Nano-Biophotonics that already are or soon will become relevant to the analysis of intracellular processes.

Simultaneous forward and epi-CARS microscopy with a single detector by time-correlated single photon counting.

We present a novel scheme to simultaneously detect coherent anti-Stokes Raman scattering (CARS) microscopy signals in the forward (F) and backward (epi - E) direction with a single avalanche photodiode (APD) detector using time-correlated single photon counting (TCSPC). By installing a mirror at a well-defined distance above the sample the forward-scattered F-CARS signal is reflected back into the microscope objective leading to spatial overlap of the F and E-CARS signals. Due to traveling an additional distance the F-CARS signal is time delayed relative to the E-CARS signal.

Pulsed-interleaved excitation FRET measurements on single duplex DNA molecules inside C-shaped nanoapertures.

Single-molecule fluorescence resonant energy transfer (FRET) is a widely accepted method for determining the spatial separation between molecules. In combination with pulsed interleaved excitation (PIE), additional information about the stoichiometry of molecular interactions is obtained. PIE-FRET, however, as implemented with standard confocal optics, requires the dilution of the sample to biologically low concentrations.