title: Effect of protease treatment on plasma infectivity in variant Creutzfeldt-Jakob disease mice.
abstract: BACKGROUND
  The emergence of variant Creutzfeldt-Jakob disease (vCJD) and of a probable transmission of the disease through blood transfusion from a presymptomatic case has underlined the need for a reliable, sensitive, and specific screening test. This study was initiated to explain why attempts to identify protease-resistant prion protein (PrPres) following treatment with proteinase K (PK) in blood or blood components have so far failed.
  
  
  METHODS
  RIII mice were inoculated intracerebrally (i.c.) with vCJD agent. As soon as some mice became ill, blood from all mice was collected, pooled, and separated into components. Aliquots of plasma were treated with either 100 and 500 microg per mL PK or left untreated. Samples were analyzed for total protein and for PrPres by Western blot with 6H4 antibodies. Infectivity in PK-treated and untreated samples was bioassayed by i.c. inoculation into healthy mice.
  
  
  RESULTS
  Estimated infectivity in untreated control plasma was 20.6 IU per mL. Treatment of plasma with 100 or 500 microg per mL PK resulted in estimated infectivity levels of 8.4 and 5.2 IU per mL, respectively. Coomassie staining revealed substantial changes in the protein profile after PK treatment, with massive degradation of proteins at 500 microg per mL PK. No PrPres was detected in plasma samples by Western blotting.
  
  
  CONCLUSIONS
  Infectivity in plasma of vCJD-infected mice showed a trend toward reduction following enzymatic treatment with increasing doses of PK, possibly because of activity against proteolysis-sensitive isoforms of abnormal prion protein. It is concluded that the use of PK in protocols for the detection of PrPres may decrease the sensitivity of blood-based assays.
authors.affiliation: Jerome H. Holland Laboratory for the Biomedical Sciences, American Red Cross, Rockville, Maryland 20855, USA.
authors.forename: Oksana
authors.initials: O
authors.lastname: Yakovleva
authors.forename: Anna
authors.initials: A
authors.lastname: Janiak
authors.forename: Carroll
authors.initials: C
authors.lastname: McKenzie
authors.forename: Lisa
authors.initials: L
authors.lastname: McShane
authors.forename: Paul
authors.initials: P
authors.lastname: Brown
authors.forename: Larisa
authors.initials: L
authors.lastname: Cervenakova
mesh_terms: D000818:Animals; D001798:Blood Proteins / Q000378:metabolism; D007562:Creutzfeldt-Jakob Syndrome / Q000097:blood*; D019286:Endopeptidase K / Q000494:pharmacology*; D051379:Mice; D018621:PrPC Proteins / Q000378:metabolism*; D013997:Time Factors
chemical_list: D001798:Blood Proteins; D018621:PrPC Proteins; D019286:Endopeptidase K