It is proposed to purchase a scanning laser confocal microscope with two-photon capabilities to measure the 3D conformation of living cells, tissues, macromolecular assemblies, and novel materials. With this new microscope, one can (i) slice clean, thin optical sections out of thick fluorescent specimens and light scattering materials, (ii) view specimen planes parallel to the line of sight, (iii) penetrate deep into light-scattering tissues, living cells, complex fluids, and complex materials, and (iv) gain 3-D views at very high resolution. This new microscope will compliment the experimental techniques currently used by Hopkins faculty, active in research projects in biomedical engineering, chemical engineering, materials science and biology. This proposal details four projects where the new SLCM will have a dramatic impact. These projects include: (1) the development of a high resolution particle-tracking instrument coupled to the new confocal microscope to probe the in vitro and in vivo mechanical properties of biopolymers; (2) studying the lateral organization of membrane proteins in epithelial cells emphasizing the packaging of glycosylphos-phatidylinositol-anchored proteins, GPI-proteins, for targeted delivery to the apical surface of morphologically polarized cells; (3) studying the molecular mechanisms by which chromatin insulators affect enhancer-promoter interactions; in particular, studying the gypsy insulator, a sequence located in the 5' transcribed untranslated region of the gypsy retrotransposon of Drosophila; (4) monitoring the process of transfecting cells and micro-organelles translocations with high spatial and time resolution. One of the primary educational objectives for acquiring a confocal microscope is to train Hopkins scientists and engineers to use one of the most versatile and promising tools of investigation in the life and materials sciences. Of course, such a powerful instrument cannot be used properly and to its full potential with out proper training. Therefore, two of the Co- PIS will be teaching a new course entitled "Practical Confocal Microscopy" to be offered during the January intersession to advanced graduate students in science and engineering who are interested in learning how to use the confocal microscope in their graduate research.
