P56
Platelet Screening by Bactalert Prior to Issue ‐ Limited Sensitivity Revealed by Residual Rate of Contamination in Outdated Screened Units.

Abstract

To date all of the existing monoclonal antibodies which react with human prion protein have been produced following immunisation with recombinant PrP, PrP from other species or synthetic peptide fragments. The aim of this project was to produce monoclonal antibodies raised against the native form of PrP c purified from human platelets. Following immunization of PrP -/-Edinburgh mice and the use of conventional hybridoma techniques a panel of 12 monoclonal antibodies has been established. These antibodies have been characterized by epitope mapping; binding to both native/denatured platelet PrP and native/denatured a-helical/b-sheet recombinant mouse PrP; immunoprecipitation of PrP c , PrP sc and PrP res from neurological control/vCJD brain homogenates and as probes for Western blotting, immunohistochemistry and flow-cytometry studies. A range of epitopes was recognised by the panel, with discrimination between normal a-helical and abnormal b-sheet forms. Several of the antibodies specifically captured abnormal forms of prion protein (PrP sc and PrP res ) found in vCJD-infected CNS and lymphoreticular tissues. It is hoped that these antibodies will aid in the early diagnosis and classification of human prion disease and the detection of vCJD infectivity in tissues for transplant and blood donations.There are presently 161 reported cases of vCJD in the UK, with three cases associated with the receipt of blood components. However, there is currently no routine large-scale screening test for vCJD in humans. The NHSBT considers that it is essential to be in a position to validate emerging tests for vCJD as quickly as possible, to ensure that suitable tests can be implemented in a timely manner, if required by the DoH. NHSBT is proactively preparing a panel of samples that will be used in assessing tests, to determine if they are operationally fit for purpose. This work is being undertaken in a specifically designed Test Assessment Facility (TAF) at Manchester Blood Centre, which went live in November 2005. To manage the panel, the Thermo Nautilus Laboratory Information Management System (LIMS) has been installed. Nautilus is a highly configurable LIMS, capable of handling large amounts of data. Irreversible anonymisation, integration with robotic equipment, and the ability to interrogate other data systems and retrieve information, are key features. Whole blood donations are received into the TAF and are logged into the LIMS, which retrieves basic donor demographic information, then irreversibly anonymises it. Red cell, buffy coat and plasma components are then prepared, and robot sample handlers sub-aliquot them. The LIMS then references a data output file, and processes the sub-aliquot information, creating a detailed inventory. Wireless technology is used for plate storage. The LIMS underwent extensive validation, including the use of designated test scripts to challenge and validate all aspects of the system, plus verification of compliance with GMP and GAMP regulations. The panel builds at a rate of around 125 don...