Cryopreservatives like dimethyl sulfoxide and glycerol are common agents that prevent cellular damage upon freezing of tissues or entire organisms. Although the cryopreservation capabilities of these compounds have been known empirically for years, much is unknown about the actual perfusion and distribution of the agents within cells on the microscopic scale. In this contribution, we report on studies that aim to uncover the dynamic distribution of cryopreservatives in the tissue with the aid of stimulated Raman scattering microscopy, enabling a direct and real-time view of the cellular loading and accumulation dynamics of these agents at the micrometer scale.
We have measured the absorption spectrum of the laser dye IR125 (also known as indocyanine green) at the water/air interface using resonant enhanced surface second harmonic generation. The spectra of the dye molecules at the interface are extremely sensitive to the bulk concentration. All the surface spectra reflect aggregation of the dye at the interface, even at the smallest concentrations detectable, below 1 (mu) M. Resonant enhanced second harmonic generation appears to be a good technique for measuring spectra at liquid interfaces.
We have made ultrafast time resolved pump probe measurements on the intramolecular electron transfer (ET) of the betaines, specifically betaine3O and penta-t-butyl betaine. The data have been analyzed to determine the ET rate in a range of solvent environments, at various temperatures and at a variety of pump and probe wavelengths. In all cases, the observed ET rate is fast, often faster than predicted by common ET theories. As a result, we have extended some common theories to successfully predict the measured ET rates. In addition to the ET dynamics, the data also display evidence for local heat deposition in the immediate vicinity of the betaine molecule which our extended model qualitatively predicts.
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