Abstract
The Ames test is designed to detect the mutagenicity of a compound. It has, however, a number of inherent flaws resulting in many compounds that are falsely reported as positives or negatives. The RIT Proteomics Lab is developing a method, utilizing two-dimensional gel electrophoresis, which will be complementary to the Ames test. Potential carcinogenicity of various compounds will be determined through analysis of the protein expression of Pseudomonas putida KT2440. Clear separation of protein spots on polyacrylamide gels is necessary to perform accurate proteomic analysis using Phoretix software. The goal of this project is to increase resolution and reproducibility of two-dimensional electrophoretic gels through a variation of multiple parameters specifically for the rehydration step. Active and passive rehydration - two common rehydration techniques for isoelectric focusing of Immobilized pH Gradient (IPG) strips - are evaluated and analyzed in replicate sets. Finally, a direct comparison is made between both methods for several pre-determined run-times.
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