Comparison of methods for preparation of 125I brachytherapy source cores for the treatment of eye cancer.

Two procedures for fixing the 125I activity on different matrices were explored. In the first method, anodic deposition of 125I on silver wire was used. In the second, 125I was adsorbed as iodate (IO3-) on alumina microspheres by using a solid-solution interface technique. While the 125I uptake was approximately 84% on the silver wires by electrodeposition method, the microspheres showed approximately 97% by the solid-solution interface technique. The low leachable (0.05%) wires, and comparatively high leachable (approximately 4%) spheres were trial encapsulated and laser welded. The encapsulated sources were measured for source strength and were found to be suitable for the treatment of eye cancer.

A holistic approach for protein secondary structure estimation from infrared spectra in H(2)O solutions.

We present an improved technique for estimating protein secondary structure content from amide I and amide III band infrared spectra. This technique combines the superposition of reference spectra of pure secondary structure elements with simultaneous aromatic side chain, water vapor, and solvent background subtraction. Previous attempts to generate structural reference spectra from a basis set of reference protein spectra have had limited success because of inaccuracies arising from sequential background subtractions and spectral normalization, arbitrary spectral band truncation, and attempted resolution of spectroscopically degenerate structure classes. We eliminated these inaccuracies by defining a single mathematical function for protein spectra, permitting all subtractions, normalizations, and amide band deconvolution steps to be performed simultaneously using a single optimization algorithm. This approach circumvents many of the problems associated with the sequential nature of previous methods, especially with regard to removing the subjectivity involved in each processing step. A key element of this technique was the calculation of reference spectra for ordered helix, unordered helix, sheet, turns, and unordered structures from a basis set of spectra of well-characterized proteins. Structural reference spectra were generated in the amide I and amide III bands, both of which have been shown to be sensitive to protein secondary structure content. We accurately account for overlaps between amide and nonamide regions and allow different structure types to have different extinction coefficients. The agreement between our structure estimates, for proteins both inside and outside the basis set, and the corresponding determinations from X-ray crystallography is good.

A holistic approach to protein secondary structure characterization using amide I band Raman spectroscopy.

We have developed a holistic protein structure estimation technique using amide I band Raman spectroscopy. This technique combines the superposition of reference spectra for pure secondary structure elements with simultaneous aromatic, fluorescence, and solvent background subtraction, and is applicable to solution, suspension, and solid protein samples. A key component of this technique was the calculation of the reference spectra for ordered helix, unordered helix, and sheet, turns, and unordered structures from a series of well-characterized reference proteins. We accurately account for the overlap between the amide I and non-amide I regions and allow for different scattering efficiencies for different secondary structures. For hydrated samples, we allowed for the possibility that bound water spectra differ from the bulk water spectra. Our computed reference spectra compare well with previous experimental and theoretical results in the literature. We have demonstrated the use of these reference spectra for the estimation of secondary structures of proteins in solution, suspension, and dry solid forms. The agreement between our structure estimates and the corresponding determinations from X-ray crystallography is good.

Long-term and high-temperature storage of supercritically-processed microparticulate protein powders.

PURPOSE: The long-term and high-temperature storage of dry, micron-sized particles of lysozyme, trypsin, and insulin was investigated. Subsequent to using supercritical carbon dioxide as an antisolvent to induce their precipitation from a dimethylsulfoxide solution, protein microparticles were stored in sealed containers at -25, -15, 0, 3, 20, 22, and 60 degrees C. The purpose of this study was to investigate the suitability of supercritical antisolvent precipitation as a finishing step in protein processing. METHODS: Karl Fisher titrations were used to determine the residual moisture content of commercial and supercritically-processed protein powders. The secondary structure of the dry protein particles was determined periodically during storage using Raman spectroscopy. The proteins were also redissolved periodically in aqueous buffers and assayed spectrophotometrically for biological activity and by circular dichroism for structural conformation in solution. RESULTS: Amide I band Raman spectra indicate that the secondary structure of the protein particles, while perturbed from that of the solution state, remained constant in time, regardless of the storage temperature. The recoverable biological activity upon reconstitution for the supercritically-processed lysozyme and trypsin microparticles was also preserved and found to be independent of storage temperature. Far UV circular dichroism spectra support the bioactivity assays and further suggest that adverse structural changes, with potential to hinder renaturation upon redissolution, do not take place during storage. CONCLUSIONS: The present study suggests that protein precipitation using supercritical fluids may yield particles suitable for long-term storage at ambient conditions.