title: Highly infectious prions generated by a single round of microplate-based protein misfolding cyclic amplification.
abstract: Measurements of the presence of prions in biological tissues or fluids rely more and more on cell-free assays. Although protein misfolding cyclic amplification (PMCA) has emerged as a valuable, sensitive tool, it is currently hampered by its lack of robustness and rapidity for high-throughput purposes. Here, we made a number of improvements making it possible to amplify the maximum levels of scrapie prions in a single 48-h round and in a microplate format. The amplification rates and the infectious titer of the PMCA-formed prions appeared similar to those derived from the in vivo laboratory bioassays. This enhanced technique also amplified efficiently prions from different species, including those responsible for human variant Creutzfeldt-Jakob disease. This new format should help in developing ultrasensitive, high-throughput prion assays for cognitive, diagnostic, and therapeutic applications. IMPORTANCE The method developed here allows large-scale, fast, and reliable cell-free amplification of subinfectious levels of prions from different species. The sensitivity and rapidity achieved approach or equal those of other recently developed prion-seeded conversion assays. Our simplified assay may be amenable to high-throughput, automated purposes and serve in a complementary manner with other recently developed assays for urgently needed antemortem diagnostic tests, by using bodily fluids containing small amounts of prion infectivity. Such a combination of assays is of paramount importance to reduce the transfusion risk in the human population and to identify asymptomatic carriers of variant Creutzfeldt-Jakob disease.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Moudjou
authors.forename: Pierre
authors.initials: P
authors.lastname: Sibille
authors.forename: Guillaume
authors.initials: G
authors.lastname: Fichet
authors.forename: Fabienne
authors.initials: F
authors.lastname: Reine
authors.forename: Jérôme
authors.initials: J
authors.lastname: Chapuis
authors.forename: Laetitia
authors.initials: L
authors.lastname: Herzog
authors.forename: Emilie
authors.initials: E
authors.lastname: Jaumain
authors.forename: Florent
authors.initials: F
authors.lastname: Laferrière
authors.forename: Charles-Adrien
authors.initials: CA
authors.lastname: Richard
authors.forename: Hubert
authors.initials: H
authors.lastname: Laude
authors.forename: Olivier
authors.initials: O
authors.lastname: Andréoletti
authors.forename: Human
authors.initials: H
authors.lastname: Rezaei
authors.forename: Vincent
authors.initials: V
authors.lastname: Béringue
mesh_terms: D000818:Animals; D019411:Clinical Laboratory Techniques / Q000379:methods*; D057166:High-Throughput Screening Assays; D006801:Humans; D018620:PrPSc Proteins / Q000032:analysis*; D017096:Prion Diseases / Q000175:diagnosis*; D012680:Sensitivity and Specificity
chemical_list: D018620:PrPSc Proteins