control mice were kept in the same room and did not develop any neurological disease. The incubation periods correspond to survival times assessed according to the criteria in (25). 29. Whole brain hemispheres were fixed in buffered 10%formalin. Pieces of brain were then embedded either in paraffin for immunohistochemistry (7-m sections) or in Araldite (4-m sections) for fine morphological examination. Antibodies were a polyclonal antibody to mouse glial fibrillary acidic protein (GFAP) and a horseradish peroxidase-conjugated secondary antibody (Dako). Seven PrPres -and six PrPres ϩ brains were examined. Spongiform lesions and gliosis could not be seen in any brain region of PrPres -mice. The absence of localized PrPres deposits was confirmed by PrP immunohistochemistry. 30. Whole brain hemispheres were fixed overnight with a solution of 1% glutaraldehyde and 1% paraformaldehyde in 0.12 M phosphate buffer (pH 7.4). After 1 hour postfixation with 2% osmic acid, they were stained en bloc with uranyl acetate and embedded in Araldite. Ultrathin sections were stained with uranyl acetate and lead citrate before examination with a Philips CM10 electron microscope.

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