With genome sequencing now almost routine in application, interest has expanded to the information embedded in the DNA sequence. Recent advancements in protein sciences and the technological development in proteomics allow exploration of gene products and functions on a larger scale. The availability of the almost complete M. avium subspecies paratuberculosis genome information and improved proteomics platforms provides an opportunity for the accelerated identification of antigens suitable for improved diagnosis and possibly prevention of Johne's disease. Currently, in our laboratory, an integrated proteomics approach consisting of liquid-phase (preparative isoelectric focusing and gel elution) and solid-phase (1- and 2-dimensional electrophoresis) separation platforms are employed in the study of secreted antigens of M. avium and M. avium subspecies paratuberculosis suitable for improved diagnosis. The liquid phase separation approach allows continuous monitoring of IFNg release assay activity (BOVIGAM^TM, CSL Animal Health Ltd) of the fractionated protein samples whereas 2-dimensional electrophoresis provides high-power resolution of individual components. Amino acid sequencing and mass spectroscopy are employed in the identification of potentially active proteins. Secreted proteins were derived from "time-course" grown cultures of M. avium (2 to 10 weeks) and M. avium subspecies paratuberculosis (7 to 34 weeks) and are being analysed using the integrated approach. A number of antigens have been already identified and tested in the IFNg release assay in conjunction with recombinant antigen screening.