http://precedings.nature.com/users/60cb0bd917acb5459a7bd302fa98f478/ Documents posted by Craig Rowell Nature Publishing Group en Nature Precedings http://precedings.nature.com/images/header_logo.gif http://precedings.nature.com http://dx.doi.org/10.1038/npre.2008.2032.1 Advances in Mass-spectrometry techniques allow for the rapid processing and evaluation of complex biological mixtures such as blood/serum. These samples represent a protein rich environment as well as a sentinel monitoring system of the entire organism. The central tenet of these studies is that changes in the microenvironment of a tissue, brought about by a disease process, will lead to sufficient changes in the protein and peptide pattern of the serum, such that the differences can be accurately detected and correctly associated with a particular disease state. Using mass-spectrometry approaches we have developed techniques that allow us to compare samples from tumor-free and tumor present serum samples simultaneously to find biomarkers that indicate the presence of cancer. To examine potentially important but less abundant proteins, ultrafiltration (UF) was used to eliminate the more abundant proteins and combine this with the non-isotopic peptide tags (S-methylthioacetimidate and S-methyl thiopropionimidate) described by Beardsley and Reilley (J. Proteome Res. 2: 15-21, 2003) to differentiate our samples. Use of these mass-coded abundance tags (MCAT) allows for simultaneous evaluation of serum samples from tumor present, and tumor free animals. Using an oa time-of-flight mass-spectrometer (Q-tof) with electrospray ionization we produce high quality spectrums to screen for peptides that have only one tag. Specificity of tagging increases the likelihood that the peptide resulted from a protein unique to either the control or conditioned state. Using the ms/ms function of the Q-tof we sequence the peptide and identify the parent protein. Specifically, our lab is using UF, MCAT and the Q-tof to evaluate rat models of chemically-induced tumors. By using animal models we overcome much of the variability that may exist in human serum samples due to differences in gender, diet and cancer initiation. We have shown that these systems allow for the identification of both small molecules such as Alpha S1 casein precursor (24 kDa) as well as proteins greater than the MCO such as Fibrinogen alpha/alpha E precursor and Coagulation factor 2 (86 and 70 kDa, respectively). With positive sequence identification we can now evaluate the tumors themselves to determine if the proteins are over-expressed in the tumor vs. normal tissues. Using this method of “bottom-up” analysis provides information on the nature and composition of our samples to more rapidly identify those proteins that are unique to the tumor state of the animals. http://dx.doi.org/10.1038/npre.2008.2032.1 Thu, 03 Jul 2008 12:27:47 UTC Serum profiling and biomarker discovery of rat mammary tumors using mass-coded abundance tags (MCAT) doi:10.1038/npre.2008.2032.1 2008-07-03 Craig Rowell Nature Precedings 2008-07-03T12:27:47Z Poster Cancer http://creativecommons.org/licenses/by/3.0/