A rapid method for isolation of polyethylene wear debris from tissue.

Ultra high molecular weight polyethylene (UHMWPE) wear debris is a major cause of osteolysis and aseptic loosening of joint replacements. The particles are phagocytosed by -activated macrophages and the foreign body response is strongly affected by the size, shape, number, and nature of the wear particles generated in vivo. The size and volumetrie concentration of UHMWPE particles influence the macrophage response, with micrometer and sub-micrometer particles displaying the highest biological activities. A high performance isolation method of polyethylene wear particles from tissue was set up, no centrifugation or ultracentrifugation is applied, no particle is lost or deformed. Characterisation of the particles was determined by SEM, micro Raman spectroscopy and computerised image analysis. Materials and Methods -Joint capsule biopsy is paraffin embedded. -20 micron thick slices are placed onto thè polycarbonate (size pore 0.2 micron) filter placed onto a polypropylene holder. -Slice are de-waxed and digested with hypochlorite. -Doublé distilled water is flushed through thè filter. -Filters are analysed by micro-Raman spectroscopy. -Filters are observed by SEM at 10.OOOX. -Particles are counted and measured. Sections of the newly formed joint capsule were characterized by the presence of a fibrous layer of variable thickness, many mononuclear phagocytes and sometimes a few giant cells. The SEM images showed that the isolation processes removed cell debris and proteins from the PE particles. Wear particles appeared as: Granules : submicron rounded particles Beads : spherical particles 1-3 microns in diameter with an irregularly textured surface (Fig.4) Fibrils : longer than 3 microns (Fig.5) Shreds and Flakes The mean size of the polyethylene particles, expressed by equivalent diameter is 0.71, range 0.49-0.95 micrometers for tissue. The amount of particles ranger between 9.02 x 109 to 23.40 x 109 per cm-1 of tissue. Discussion and conclusion Visualization of individual particles by SEM greatly enhanced the morphologic description, otherwise difficult or impossible from histology. The proposed method comes from the necessity to overcome the technical pitfalls that we experimented when applying already published methods. Our method enabled us to retrieve polyethylene wear particles in a relatively quick time both from tissue slices digested directly on the filter that will be analysed by SEM. Numeric and morhological data are comparable to other methods.