Imaging and Spectroscopy of Biomaterials and Nanoparticle-enhanced Biomaterials

Faculty Mentor - Dr. Thomas M. Nordlund

Multiphoton Spectroscopy and Imaging. One of the best microscopies for determining structure and function of living cells and microstructural components (e.g., microfibers) in highly-scattering media is confocal microscopy, where a laser beam is passed through a microscope objective lens and scanned across a sample. The sample is labeled with a fluorescent dye, which glows when the laser excites it. A significant improvement in this technique was developed several years ago-- multiphoton excitation-- in which a laser beam of twice or three times the wavelength needed to excite fluorescence is focused deep into the sample. Because the wavelength is longer, the beam penetrates deeper into scattering media. Since the beam does not have the proper wavelength to excite dye fluorescence, no fluorescence is created until the beam reaches its point of tightest focus. There, nonlinear optical processes take place, effectively doubling or tripling the frequency of the laser light, allowing fluorescence excitation. During the summer of 2002 we constructed a stand-alone and a microscope-attached (Olympus IX70) sample holder, which allows for both imaging and measurement of fluorescence spectra of samples. Ti:sapphire and a 532-nm diode lasers are used for excitation. Nanoparticle Fluorescence Enhancement. Recently it has been shown that intrinsic DNA fluorescence, which is extremely weak under normal conditions, can be enhanced by a large factor if DNA is located 5-20 nm from metallic silver particle surfaces. We have learned both vapor-deposition and colloid-suspension methods of silver nanoparticle preparations. We are now characterizing SiO2 coatings on the nanoparticles, attaching linking groups to the surface, and attaching DNA to the linkers. We believe these preparations can be used to create excitations at one end of the DNA that will travel via energy transfer down the DNA helix—a molecular “fiber optic”. A Microsphere Screening Method for Sunscreen Agents. We have developed a method for adsorbing hydrophobic sunscreen agents to polystyrene microspheres, to test whether added DNA will (i) interact with the sunscreen and (ii) accept transferred UV excitation energy from the sunscreen. If the latter is observed, the sunscreen is a candidate for further testing for failing to prevent damage to DNA.

Examples of projects to be undertaken by an undergraduate at UAB:

  • ¨Simultaneously image and measure fluorescence spectra of dye-labeled cells, DNA or microspheres.
  • Measure optical spectra of DNA attached to silver nanoparticles, prepared by a graduate student. Determine whether absorption, fluorescence and energy transfer are enhanced.
  • Add sunscreen to coat microsphere surfaces; titrate with DNA, measuring fluorescence and absorption spectra, both standard and laser-excited. Fit data with available computer programs.
  • Molecular modeling of DNA and cinnamate structure. Attempt "dock" of molecules on computer.

REU students Harriet Crockett, Aaron Carr, Andreia Williams, Rosalind Ramsey, Steve Davis, and Michael Lee have participated in some of these or related projects since 1998.