We undertook to identify and express proteins of Mycobacterium avium subspecies paratuberculosis (MAP) for use in evaluating their potential as biomarkers for the presence of this organism in infected animals. Such proteins, produced synthetically, may also have potential for application as refined diagnostic antigens in enhancement of the γ-IFN assay. Using liquid-phase proteomics we fractionated the cell-free supernatants (CFS) of MAP grown in vitro and using automated N-terminal sequencing identified: Antigen D, SOD, MAP2609 and Ahp C. These four proteins, and along with Mp35K, Hsp65, Ahp D, P34 and PstS3, which were rationally-selected with reference to the Mycobacterium avium (MAV) and MAP genomes, were cloned and expressed in, then purified from E. coli. Several of these proteins were detected by antibodies present in the serum of animals experimentally-challenged with MAP; however, their utility as MAP biomarkers remains unclear.
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