Polarization-sensitive sum-frequency generation microscopy of collagen fibers.

Point-scanning sum-frequency generation (SFG) microscopy enables the generation of images of collagen I fibers in tissues by tuning into specific vibrational resonances of the polypeptide. It is shown that when collagen-rich tissues are visualized near the 2954 cm(-1) stretching vibration of methylene groups, the SFG image contrast is higher compared to the contrast seen in nonresonant second-harmonic generation (SHG) imaging. Polarization and spectrally resolved analysis of the SFG signal as a function of fiber orientation in the CH-stretching range of the vibrational spectrum enabled a comparative characterization of the achiral tensor elements of collagen's second-order susceptibility. This analysis reveals that selected on-resonance tensor elements are enhanced over other elements, giving rise to a much stronger anisotropy ρ of the signal for SFG (ρ ≈ 15) compared to SHG (ρ ≈ 3). The improved anisotropy of the vibrationally resonant signal contributes to the higher contrast seen in the SFG tissue images.

Vibrationally resonant sum-frequency generation microscopy with a solid immersion lens.

We use a hemispheric sapphire lens in combination with an off-axis parabolic mirror to demonstrate high-resolution vibrationally resonant sum-frequency generation (VR-SFG) microscopy in the mid-infrared range. With the sapphire lens as an immersed solid medium, the numerical aperture (NA) of the parabolic mirror objective is enhanced by a factor of 1.72, from 0.42 to 0.72, close to the theoretical value of 1.76 ( = nsapphire). The measured lateral resolution is as high as 0.64 µm. We show the practical utility of the sapphire immersion lens by imaging collagen-rich tissues with and without the solid immersion lens.

Controlling stimulated coherent spectroscopy and microscopy by a position-dependent phase.

We study the role of geometry dependent phase shifts in stimulated coherent spectroscopy, a special class of heterodyne optical spectroscopy techniques. We generalize the theoretical description of stimulated spectroscopy to include spatial phase effects, and consider the measured material response for several representative excitation and detection configurations. Using stimulated Raman scattering microscopy as an example, we show that different components of the material response are measured depending the position of the object in focus. We discuss the implications of the position dependent phase in stimulated coherent microscopy and point out a detection configuration in which its effects are minimized.

Dissociation chemistry of hydrogen-deficient radical peptide anions.

The fragmentation chemistry of anionic deprotonated hydrogen-deficient radical peptides is investigated. Homolytic photodissociation of carbon-iodine bonds with 266 nm light is used to generate the radical species, which are subsequently subjected to collisional activation to induce further dissociation. The charges do not play a central role in the fragmentation chemistry; hence deprotonated peptides that fragment via radical directed dissociation do so via mechanisms which have been reported previously for protonated peptides. However, charge polarity does influence the overall fragmentation of the peptide. For example, the absence of mobile protons favors radical directed dissociation for singly deprotonated peptides. Similarly, a favorable dissociation mechanism initiated at the N-terminus is more notable for anionic peptides where the N-terminus is not protonated (which inhibits the mechanism). In addition, collisional activation of the anionic peptides containing carbon-iodine bonds leads to homolytic cleavage and generation of the radical species, which is not observed for protonated peptides presumably due to competition from lower energy dissociation channels. Finally, for multiply deprotonated radical peptides, electron detachment becomes a competitive channel both during the initial photoactivation and following subsequent collisional activation of the radical. Possible mechanisms that might account for this novel collision-induced electron detachment are discussed.

Psychometric properties of the life events checklist.

The Life Events Checklist (LEC), a measure of exposure to potentially traumatic events, was developed at the National Center for Posttraumatic Stress Disorder (PTSD) concurrently with the Clinician Administered PTSD Scale (CAPS) to facilitate the diagnosis of PTSD. Although the CAPS is recognized as the gold standard in PTSD symptom assessment, the psychometric soundness of the LEC has never been formally evaluated. The studies reported here describe the performance of the LEC in two samples: college undergraduates and combat veterans. The LEC exhibited adequate temporal stability, good convergence with an established measure of trauma history -- the Traumatic Life Events Questionnaire (TLEQ) -- and was comparable to the TLEQ in associations with variables known to be correlated with traumatic exposure in a sample of undergraduates. In a clinical sample of combat veterans, the LEC was significantly correlated, in the predicted directions, with measures of psychological distress and was strongly associated with PTSD symptoms.