I. P. Lipscomb

Prion diseases or transmissible spongiform encephalopathies (TSEs) are a group of rare, transmissible and fatal neurodegenerative diseases associated with the protein agent (PrP^Sc). As such, the sensitive and rapid detection of prion PrP^Sc amyloid on the surface of suspect surgical instruments is of great importance and may even allow remedial action to be taken prior to any further operative intervention and possible iatrogenic transmission. However, conventional PrP^Sc detection methodologies tend to rely on the inefficient and unreliable removal of suspect material from a surface using swabs or wipes prior to antibody analysis. Here we show how the combination of an advanced light microscope technique, episcopic differential interference contrast/epifluorescence (EDIC/EF) microscopy, and the application of β-amyloid fluorescent thiazole markers (thioflavin T, thioflavin S) can be used to detect, in situ, submicron (attomole) levels of prion protein amyloid contamination in brain and spleen sections, smears and homogenate on surgical stainless steel surfaces and surgical instruments. This technique, although not specific to an amyloid type, can be used to verify that surgical instruments are substantially free from prion amyloid protein soiling and hence reduce the risk of iatrogenic transmission.

The ineffective cleaning of surgical instruments may be a vector for the transmission of hospital-acquired infections. The aim of this research was to investigate whether further decontamination procedures need to be instigated in sterile-service departments (SSDs) to reduce the risk of nosocomial illnesses, such as endotoxemia, sepsis, or iatrogenic Creutzfeldt-Jakob disease (to date, 1,147 cases of confirmed Creutzfeldt-Jakob disease deaths in the United Kingdom since 1990 have been reported). Instrument sets were obtained from nine anonymous United Kingdom National Health Service (NHS) primary care trust SSDs. The investigation implemented an advanced light microscopy technique, episcopic differential interference contrast microscopy with the sensitive fluorescent reagents SYPRO Ruby and 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI), to detect proteinaceous and microbial contamination levels. Gram-negative lipopolysaccharide (LPS) endotoxin was monitored using a dansylated polymyxin B fluorochrome agent. None of the 260 instruments examined displayed signs of microbial colonization or LPS endotoxin contamination. However, over 60 percent of the instruments showed a high degree of protein soiling (0.4 to 4.2 μg protein/mm ² ). Some instruments appeared soiled with crystalline deposits that may consist of a potentially hazardous material contributing to inflammation and/or surgical shock. It is clear that the overall standard for cleaning must be raised in order to fulfill the imminent introduction of new European standards and to reduce the risk of cross-patient contamination and iatrogenic transmission.