UNIVERSITY OF PITTSBURGH SCHOOL OF MEDICINE
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Division of Neuropathology University of Pittsburgh School of Medicine Room A506 PUH 200 Lothrop Street Pittsburgh, PA 15213 Tel: 412-647-9417 |
Biosafety Manual
Technical Procedures for Decontaminating Tissues and Inactivating
Suspected Creutzfeldt-Jakob Disease
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For further information see WHO Infection Control Guidelines for Transmissible Spongiform Encephalopathies Creutzfeldt-Jakob disease is a progressive fatal disease of the central nervous system, which occurs in middle life, affecting both sexes. After a period of vague prodromal symptoms, dementia appears, and progresses rapidly to coma and death, usually within one to two years of onset, though survival for more than five years has been reported. Dementia is accompanied by a variety of other neurologic disturbances; there may be clinical evidence of focal critical degeneration, upper and lower motor-neuron signs, extrapyramidal signs, pareses, cerebellar ataxia and seizures. Myoclonus is particularly characteristic which is an irregular involuntary contraction of a muscle usually resulting from functional disorder of controlling motor neurons. Creutzfeldt-Jakob disease has been observed in combination with other neurologic disorders, including multiple sclerosis, cerebrovascular accidents, vasculitis, tumors, hepatocellular degeneration, post-traumatic syndromes and Alzheimer's disease. Because of the great variability in clinical findings, any adult who acquires rapidly progressive dementia without having a spacing-occupying intracranial lesion should be suspected of having Creutzfeldt-Jakob disease; the presence of myoclonus should greatly increase this suspicion. The natural modes of transmission of the agent of the disease are unknown. Any break in the body's integument probably constitutes a portal of entry for prions. This is why universal precautions should always be practiced. Equipment for Recommended Decomtamination Procedures
Ineffective Procedures The prion resists inactivation by: boiling, ultraviolet irradiation, ethylene oxide sterilization, ethanol, formalin, beta-propiolactone, detergents, quaternary ammonium compounds, Lysol, alcoholic iodine, acetone, potassium permanganate. The problem with many of the above stated procedures is unfixed tissue does not withstand any of those treatments, formalin fixation stabilizes ineffectively against the sterilization action of the autoclave and exposure of fixed tissue to either sodium hydroxide or bleach produces unacceptable histologic artifacts. Formic acid inactivates the prion in unfixed brain and is used to enhance immunostaining. Exposing formalin-fixed tissue to formic-acid (>-95%) is an excellent alternative to the formalin-phenol fixation. Tissue preparation procedure prior to processing:
These procedures will sterilize CJD-contaminated tissue and materials and must be followed for autopsy instruments, specimen containers and their solutions and other laboratory instruments. |
Technical Procedures For Autopsy And Surgical Tissue Specimens
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Autopsy: The Infectious Room is used in the Morgue. Technical personnel should wear gloves, gown or jumpsuit, shoe covers, mask and face shield. Disposable instruments are used if possible i.e. brain knife and scalpel. All instruments, cassettes, containers, etc need to be set up prior to the Neuropathologist cutting the brain. The table is lined with freezer paper. As the neuropathologist cuts the brain the technician tissue cassettes with the corresponding accession number and area of the brain the section came from. The tissue cassettes are then placed in a disposable specimen bottle containing formalin, if sections are not adequately fixed. After all sections are submitted, the specimen container is placed in a zip lock bag and placed in a blue biohazard carry bag to be transported to the laboratory. The large bucket containing the CJ infected brain is placed inside a red biohazard bag and secured. The freezer paper is disposed of in the trash can lined with a red biohazard bag. A solution of 1 Normal Sodium Hydroxide is poured onto the instruments, tables, etc (anything else that came in contact with the brain) and allowed to soak for 1 hour to insure inactivation. A "test" block fixed in formalin overnight and for 2 hours in formic acid of the suspected CJD infected case is submitted prior to the entire case being deactivated and processed. This test block will give the pathologist a preliminary diagnosis as to whether the case is the positive for CJ disease. If the case is positive, the following procedure needs to be followed. Once the fixed tissue blocks are transported to the laboratory they are placed in formic acid > 95% for 2 hours to deactivate the agent. The sections are washed well in tap water then placed back into 10% formalin for a minimum of 2 days prior to processing on tissue processor. If a surgical biopsy is performed and suspected of Creutzfeldt Jakob disease the tissue must be fixed well in formalin at least 24 hours/overnight fixation, then placed in 95-100% formic acid for 30-60 minutes and placed back into formalin for at least 2 hours.
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