The complexity of current problems in biophysics and cell biology increasing require chemical and physical-based methods to glean meaningful solutions. Central to this approach is the development of new technologies. Research efforts in my lab are directed at further developing nonlinear optical spectroscopy and microscopy methods to address problems related to structural protein arrays (e.g. collagen, and acto-myosin complexes). One long-term goal is to disseminate new tools for high resolution clinical imaging applications that are more powerful than current methods. We are also developing new optical methods for nano/microfabrication of tissue engineering scaffolds. The group currently works on the following three projects:
1) Second Harmonic Generation (SHG) imaging of tissues
We are developing SHG imaging microscopy as an “optical biopsy” tool to differentiate normal and diseases states. Since SHG is highly sensitive to the supramolecular structure of structural protein arrays, this imaging modality is a powerful approach to this problem. A central theme of this project is to develop a thorough understanding of the underlying chemical and physical properties that give rise to the SHG. This information then guides us to the appropriate optical signatures in assessing diseased tissues. We are currently focusing on muscular and skeletal disorders including aging and Osteogenesis Imperfecta, respectively. Through quantitative SHG measurements, we have observed that this technique provides clear delineation in these pathologies, and suggests its use as a clinical diagnostic tool.
2) Multiphoton excited (MPE) nano/microfabrication of tissue engineering scaffolds
A major challenge in tissue engineering is the fabrication of multiscale (i.e. Extra Cellular Matrix-cell-tissue) scaffolds that provide the topographic and biochemical ECM cues found in native tissue such as skin. To solve this problem, we use MPE photochemistry to fabricate synthetic ECMs directly from structural proteins including collagen, fibronectin, and laminin. The approach is analogous to that of two-photon excited fluorescence microscopy, but rather than imaging plane by plane, we exploit the multiphoton confinement of the excitation to the focal plane to nano/microfabricate 3-D structures. The project entails development of new optical instrumentation to replicate the complex structure of the native ECM, physical and chemical characterization of the scaffolds, as well as developing a fundamental understanding of the topographic and biochemical factors that are responsible for cell differentiation and new tissue synthesis.
3) MPE Intracellular Nanofabrication
A new project uses multiphoton excited photochemistry to fabricate structures inside live cells as a means of studying spatially localized signaling events. We can fabricate walls, channels, and corridors directly from cytoplasmic proteins. Our results indicate that these structures form effective barriers to diffusion. In collaboration with Dr. Vladimer Rodionov’s group, we will use this method to examine microtubule dynamics in fibroblasts.
