pkh-26 and Transfusion-Reaction

Contamination of blood products by white blood cells leads to a risk of transmission of infectious agents, particularly abnormal prion protein, the probable causative agent of new-variant Creutzfeldt-Jakob disease. Blood product filtration could reduce this risk, but the filtration systems might generate potentially infectious membrane fragments. We developed an original flow cytometric method that allows the detection and quantification of membrane fragments in filtered products and the evaluation of the quantity of destroyed cells.. This method has four technical requirements: cytofluorometric acquisition of forward scatter parameters on a log scale, use of a fluorescent aliphatic reporter molecule (PKH26-GL) to identify membrane fragments, quantification with fluorescent beads, and the drawing up of a standard curve on the basis of cells destroyed by freezing/thawing to generate cell debris (i.e., quantity of membrane fragments measured versus quantity of destroyed cells).. This original method can be used to test new filtration devices and it allows optimization of the filtration process or comparison of different filtration systems. We tested the method with three commercial white cell removal filters. We demonstrated that it is possible to evaluate the filter quality, particularly the likelihood of fragment removal during the filtration process.

Topics: Blood Component Removal; Blood Transfusion; Cell Membrane Structures; Cell Separation; Creutzfeldt-Jakob Syndrome; Filtration; Flow Cytometry; Fluorescent Dyes; Humans; Leukocytes; Light; Organic Chemicals; Safety; Scattering, Radiation; Subcellular Fractions; Transfusion Reaction