Our research focuses on optical and calorimetric analytical instrumentation applicable to fundamental problems in the life sciences.
The technique of fluorescence photobleaching recovery permits the lateral motions of molecules in the membranes of single, living cells to be examined under physiological conditions. Crosslinking of membrane receptors reduces the lateral mobility of the receptors as measured by FPR. Ongoing FPR studies of white blood cell stimulation by antigens and by plant lectins are showing what specific membrane crosslinking events initiate various types of immunological processes in cells. This work also involves fluorescence-activated flow cytometry. Cell sorters allow us to measure the binding constants of substances to cell surfaces and to determine which populations of cells respond to various stimuli.
Two related experimental techniques, fluorescence depletion anisotropy and time-resolved phosphorescence anisotropy, are being used to examine rotation of macromolecules in viscous environments. Such rotations are especially sensitive to molecular aggregation state and asymmetry. We are using these methods to see, for example, how allergens trigger allergic responses in basophils and mast cells by aggregating IgE receptors. These methods are also important in the studies of immune processes described above.
Computerized image processing is another powerful tool in cell surface studies. By combining ultra low light video microscopy with image analysis computers, we can trace single fluorescent molecules moving on cell surfaces. Studies of the distribution and motion of luteinizing hormone receptors on sheep luteal cells help clarify the mechanisms by which peptide hormones regulate and maintain pregnancy.
Our other principal area of interest is microcalorimetry. Small scale determination of heats of chemical reactions and biological processes are being used in several ongoing projects. Toxicity of chemicals to cultured mammalian cells is being evaluated by flow microcalorimetry. Thermodynamics of substrate and inhibitor binding to the enzyme pyruvate kinase is under study by isoperibol titration calorimetry. Finally, data analysis methods for differential scanning calorimetry are being developed to resolve melting parameters for multidomain macromolecules like immunoglobulins and tRNAs.