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Equine protozoal myeloencephalitis (EPM) is a common and costly neurologic disease of horses caused by Sarcocystis neurona, protozoa that parasitize the nervous system. Confirmation of the clinical diagnosis is a challenge for veterinarians because the current immunologic tests lack specificity. Multiplexing strategy approaches including two-dimensional fluorescence difference gel electrophoresis (2D DIGE) and mass spectrometry were employed to identify differential protein expression associated with EPM.
Cerebrospinal fluid (CSF) samples were collected from normal and EPM horses. EPM was diagnosed if neurological signs were present, Western blot against S. neurona on the CSF was positive, and other neurological diseases were excluded by ancillary tests and/or necropsy. CSF was collected under general anesthesia, at the atlanto-occipital site to avoid blood contamination. The CSF proteins were precipitated, labeled with CyDye DIGE fluor dyes, and separated on 2D gels. The 2D gels were scanned with Typhoon, and the digitized images were analyzed with DeCyder software. The differentially expressed proteins were excised with a ProPic II spot picker, in-gel trypsin digested, and the tryptic peptides were analyzed with the electrospray ionization mass spectrometer.
A number of differentially expressed proteins were identified in the various horse CSF, including serum albumin, haptoglobin, prostaglandin-D synthase, and apolipo-proteins E and A-I. Various isoforms of serum albumin were up-regulated with EPM compared to the normal horse CSF. The above proteins, including the isoforms, may be associated with EPM, and the characterization of differentially expressed additional proteins could give insight into the pathogenesis and diagnosis of EPM.