DETECTION OF REGIONAL DIFFERENCES IN PROTEIN EXPRESSION IN HUMAN BRAIN TISSUE USING ANTIBODY MICROARRAYS

C.A. CaamaƱo; E.G. Jones; W.E. Bunney; R.M. Myers; S.J. Watson; H. Akil
Society for Neuroscience 33rd Annual Meeting. 2003.

Abstract

As a first step towards the study of protein function and regulation, we explored the use of an antibody microarray approach for the analysis of differential protein expression in human brain. We initially focused in protocol standardizations using two brain regions, dorsolateral prefrontal cortex (DLPFC) and cerebellar cortex (CB), which have been implicated in the dysregulation of gene expression in human affective disorders. Protein extracts from these two regions were obtained under non-denaturing conditions, labeled with either Cy3 or Cy5 fluorophores, mixed, and incubated with a BD-Clontech antibody microarray, containing 379 distinct monoclonal antibodies. To reduce assay variance, measurements were internally normalized by including a second array incubated with the same extracts labeled in a reverse order. Expression differences between DLPFC and CB occurred in proteins spanning broad functional and cellular localization spectra. These proteins are being investigated by Western immunoblotting to address antibody specificity and also to provide an independent estimate of their relative expression levels.Support Contributed By: The Pritzker Neuropsychiatric Disorder Research Consortium Fund and NIH Conte grant MH60398-03. We acknowledge participation of all Consortium members in this work.