Decontamination of prions in a plasma product manufacturing environment.

BACKGROUND: The high resistance of prions to inactivating treatments requires the proper management of decontaminating procedures of equipment in contact with materials of human or animal origin destined for medical purposes. Sodium hydroxide (NaOH) is widely used today for this purpose as it inactivates a wide variety of pathogens including prions. STUDY DESIGN AND METHODS: Several NaOH treatments were tested on prions bound to either stainless steel or chromatographic resins in industrial conditions with multiple prion strains. RESULTS: Data show a strong correlation between inactivation results obtained by immunochemical detection of the prion protein and those obtained with infectivity assays and establish effective inactivation treatments for prions bound to stainless steel or chromatographic resins (ion exchange and affinity), including treatments with lower NaOH concentrations. Furthermore, no obvious strain-specific behavior difference was observed between experimental models. CONCLUSION: The results generated by these investigations show that industrial NaOH decontamination regimens (in combination with the NaCl elution in the case of the chromatography process) attain substantial prion inactivation and/or removal between batches, thus providing added assurance to the biologic safety of the final plasma-derived medicinal products.

Comparison of nanofiltration efficacy in reducing infectivity of centrifuged versus ultracentrifuged 263K scrapie-infected brain homogenates in "spiked" albumin solutions.

BACKGROUND: The safety of plasma-derived products is of concern for possible transmission of variant Creutzfeldt-Jakob disease. The absence of validated screening tests requires the use of procedures to remove or inactivate prions during the manufacture of plasma-derived products to minimize the risk of transmission. These procedures need proper validation studies based on spiking human plasma or intermediate fractions of plasma fractionation with prions in a form as close as possible to that present in blood. STUDY DESIGN AND METHODS: Human albumin was spiked with low-speed or high-speed supernatants of 263K scrapie-infected hamster brain homogenates. Spiked albumin was then passed through a cascade of filters from 100 nm down to 20 to 15 nm. Residual infectivity was measured by bioassay. RESULTS: The overall removal of infectivity spiked into albumin through serial nanofiltration steps was 4 to 5 logs using low-speed supernatant and 2 to 3 logs with high-speed supernatant. CONCLUSION: These findings confirm the utility of nanofiltration in removing infectivity from plasma (or other products) spiked with scrapie brain homogenate supernatants. However, efficiency is diminished using supernatants that have been ultracentrifuged to reduce aggregated forms of the infectious agent. Thus, filtration removal data based on experiments using "standard" low-speed centrifugation supernatants might overestimate the amount of prion removal in plasma or urine-derived therapeutic products.

In vitro infectivity assay for prion titration for application to the evaluation of the prion removal capacity of biological products manufacturing processes.

Prions can be detected and quantified currently by using either immunoassays such as Western-blot, ELISA or conformation dependent immunoassay, or an infectivity assay in laboratory animals (bioassay). While immunoassays are inexpensive and rapid, they are based on the detection of PrP(Sc), the abnormal isoform of the prion protein, a surrogate marker for prion infectivity. The bioassay is considered the gold-standard analytical method for measuring prion infectivity, but it is very costly and time-consuming, involving the destruction of large numbers of animals. The use of the transgenic MovS6 cell line is described for the development of an in vitro tissue culture infectivity assay (TCIA) for prion detection and quantitation. Compared to a bioassay, the TCIA is rapid ( approximately 8 weeks), easy to implement, much less expensive, and requires far fewer animals. After titrating concomitantly a prion-infected brain homogenate sample by Western-blot, TCIA and bioassay, data show that the sensitivity of the TCIA is close to that of the bioassay, since 1 TCID(50) corresponds to 4 ID(50), and 80-fold more sensitive than the Western-blot. The application of the TCIA to the evaluation of prion removal in biological products manufacturing processes is described using a 15 nm-nanofiltration step of human albumin as a model.