AbstractThe presence of abnormal, disease-related prion protein (PrP^D) has recently been demonstrated by protein misfolding cyclic amplification (PMCA) in urine of patients affected with variant Creutzfeldt-Jakob disease (vCJD), a prion disease typically acquired from consumption of prion contaminated bovine meat. The complexity and multistage process of urine excretion along with the obligatory use of PMCA raise the issue of whether strain characteristics of the PrP^D present in vCJD brains, such as infectivity and phenotype determination, are maintained in urine excreted PrP^D and following amplification by PMCA. We inoculated transgenic mice expressing normal human PrP with amplified urine and brain homogenate achieving the same 100% attack rate, similar incubation periods (in both cases extremely long) and histopathological features as for type and severity of the lesions. Furthermore, PrP^D characteristics analyzed by immunoblot and conformational stability immunoassay were indistinguishable. Inoculation of raw vCJD urine caused no disease, confirming the extremely low concentration of PrP^D in vCJD urine. These findings show that strain characteristics of vCJD brain PrP^D, including infectivity, are preserved in PrP^D present in urine and are faithfully amplified by means of PMCA; moreover, they suggest that the PrP^D urine test might allow for the diagnosis and identification of disease subtype also in sporadic CJD.