Olive Drummond

Application of a Time–Resolved Fluoroimmunoassay for the Analysis of Normal Prion Protein in Human Blood and Its Components

Background and Objectives: To quantify the cellular isoform of prion protein (PrPc) in human blood using a new time‐resolved dissociation‐enhanced fluoroimmunoas‐say (DELFIA®). Materials and Methods: The DELFIA was optimised for human blood samples and applied to isolated cell and plasma fractions from blood donations. The physicochemical properties of PrPc were analysed. Results: 26.5% of blood PrPc was associated with the platelet fraction, 0.8% with polymorphonuclear leucocytes, 2.4% with mononuclear leucocytes, 1.8% with red cells and 68.5% with plasma (mean values from 4 processed donations). Conclusion: The majority of blood PrPc is found in the platelet and plasma compartments.

The effect of methylene blue photoinactivation and methylene blue removal on the quality of fresh-frozen plasma

BACKGROUND: T he effects of using fresh or frozen‐thawed plasma, WBC reduction of plasma before freezing, and the use of two different methylene blue (MB) removal filters on the quality of MB‐treated plasma were compared.

A potentially improved approach to methylene blue virus inactivation of plasma: the Maco Pharma Maco-Tronic system

. Plasma was subjected to methylene blue (MB) photochemical virus inactivation using the Maco Pharma Maco‐Tronic system which allows three units to be illuminated together, thus reducing processing time. The plasma bag system used incorporates an integral membrane plasma filter and a dry MB pill which dissolves in the plasma to give a 1‐µm concentration. There is computer‐controlled processing and datalogging.

The effects of leukodepletion on the generation and removal of microvesicles and prion protein in blood components.

BACKGROUND:  Universal leukodepletion (LD) has been implemented in the United Kingdom to reduce the risk of transfusion‐transmitted variant Creutzfeldt‐Jakob disease. If LD causes microvesiculation of blood cells, however, potentially infectious membrane‐associated prion could reach the final products.

Pathogen reduction of fresh plasma using riboflavin and ultraviolet light: effects on plasma coagulation proteins.

BACKGROUND: Treatment with riboflavin and ultraviolet (UV) light reduces the pathogens present in blood components. This study assessed changes to the coagulation proteins that had occurred during this treatment of fresh plasma units before freezing.

The expression of prion protein by endothelial cells: a source of the plasma form of prion protein?

Summary. The neuronal prion protein (PrPC) is also expressed within peripheral tissues including human blood. The majority of blood PrPC is found within the plasma fraction. We hypothesized that the vascular endothelium could be a source of this PrPC. Reverse transcription polymerase chain reaction demonstrated that both human umbilical vein endothelial cells (HUVEC) and human microvascular endothelial cells (HMEC‐1) expressed PrPC mRNA. Flow cytometry confirmed PrPC expression on HMEC‐1s and HUVECs (120 900 ± 15 058 and 58 327 ± 4577 molecules PrPC/cell respectively), with no upregulation following cellular activation. Confocal immunofluorescence microscopy confirmed that HMEC‐1s and HUVECs were positive for PrPC on the plasma membrane. Time‐resolved dissociation‐enhanced fluoroimmunoassay (DELFIA®) analysis of cell culture medium demonstrated a slow constitutive release of soluble PrPC not associated with activation. In contrast to von Willebrand factor antigen, PrPC plasma levels in vivo decrease following desmopressin therapy in patients with von Willebrand disease. Measurement of PrPC plasma levels in patients with varying blood counts demonstrated no association between cell count and PrPC concentration. However, there was a higher level of PrPC in plasma from patients with end‐stage renal failure. In conclusion, endothelial cells of both macrovascular and microvascular origin expressed high levels of PrPC which can be constitutively released into the cell culture medium.

Cold storage of pooled, buffy‐coat‐derived, leucoreduced platelets in plasma

Background  This study was designed to determine which in vitro assays would be most useful for studying the effects of cold storage on platelet concentrates and to establish an in vivo model for platelet recovery and survival.

The release of prion protein from platelets during storage of apheresis platelets

BACKGROUND: Recent studies using a time‐resolved fluoroimmunoassay method (dissociation‐enhanced lanthanide fluoroimmunoassay) showed that platelets and plasma are the main reservoir of the normal isoform of cell‐associated prion protein (PrPc) in human blood. The aims of the present study were to monitor PrPc levels in various fractions of apheresis platelets during storage by using the DELFIA method and to assess the association of this release with alpha‐granule protein β‐thrombo‐globulin and cytoplasmic LDH.

Freezing of buffy coat–derived, leukoreduced platelet concentrates in 6 percent dimethyl sulfoxide

BACKGROUND: There has recently been renewed interest in freezing platelets (PLTs) in dimethyl sulfoxide (DMSO) for the treatment of major traumatic injuries, especially in military situations. This study examined PLTs that were frozen in small volumes of 6 percent DMSO at −80°C.

Species differences in the blood content of the normal cellular isoform of prion protein, PrPc, measured by time-resolved fluoroimmunoassay

Background and Objectives  The concern that variant Creutzfeldt‐Jakob disease could be transmitted via blood transfusion has prompted studies of blood infectivity in animal models. As normal prion protein acts as a substrate for conversion to the abnormal form associated with infectivity, we have quantified its distribution in mice and hamsters, the most commonly used animal models.