A technique for detecting prions in tissue, developed in recent years by UCSF scientists, is significantly more sensitive than the diagnostic procedures currently used to detect the lethal particles in samples of brain tissue from patients, according to a study performed by a UCSF team.
The finding indicates that the diagnostic technique, known as the conformation-dependent immunoassay (CDI), should be established as the standard approach for brain biopsies of patients suspected of having the disease, they say. The team is exploring whether the CDI might be adapted to detect prions in blood and muscle.
The finding suggests that reliance on the current methods for detecting prions in human brain tissue -- microscopic examination of tissue for the telltale vacuoles that form in brain cells and immunohistochemistry (IHC), which involves detecting prions in brain sections using prion protein-specific antibodies -- may have led to an under diagnosis of the disease in patients in recent years, they say. (A definitive diagnosis of the disease in humans is made only on autopsy, when a neuropathologist can analyze multiple brain regions for vacuoles and evidence of prions by IHC, and it is estimated that only 50 percent of human cases are autopsied, in part because many pathologists do not want to risk infection during the autopsy.)
In the study, the team compared the ability of the CDI and the two traditional diagnostic techniques to detect prions in various brain samples from 28 patients diagnosed on autopsy as having one of several human forms of the disease -- sporadic, familial or iatrogenic Creutzfeldt-Jakob disease (CJD). While the CDI detected the biochemical signal for prions in 100 percent of the samples studied, the traditional tests failed to detect the prion in a high proportion of cases. For example, in an experiment that focused on 18 brain regions from eight patients with sporadic CJD, the CDI detected prions in 100 percent of the samples, while IHC detected them in 22 percent and routine tissue examination in 17 percent.
"In about 80 percent of the different brain regions examined, prions were not consistently detected by either IHC or routine histology that measure vacuolation. In contrast, the CDI was always positive in all regions of the brain," says the lead author of the study, Jiri Safar, MD, associate adjunct professor of neurology and a member of the UCSF Institute for Neurodegenerative Diseases, which is directed by senior author Stanley B. Prusiner, MD, UCSF professor of neurology and biochemistry.
"These findings indicate that histology and immunohistochemistry should no longer be used to rule out prion disease in single-site biopsy samples," says Safar. "The superior performance of the CDI in diagnosing prion disease suggests that the CDI be used in future diagnostic evaluations of prion disease, particularly for single-site brain biopsies during life"
"If the traditional techniques are used at autopsy, they must be applied to many cortical and subcortical samples," says co-author Stephen J. DeArmond, MD, PhD, UCSF professor of neuropathology.
Moreover, while the study examined the efficacy of the CDI in comparison to the two techniques routinely used by neuropathologists to detect prions in human brain tissue, previous studies at UCSF indicate that the CDI is also significantly more sensitive than Western blot analysis, the technology used with IHC to detect prions in brain tissue from cattle suspected of having bovine spongiform encephalopathy (BSE). That IHC and Western blot analysis are relatively insensitive methods, the researchers say, supports their ongoing assertion that the CDI should also be used to evaluate the brain tissue of cattle.
"The studies reported here are likely to change profoundly the approach to the diagnosis of prion disease in both humans and livestock," says Safar.
More broadly, the scientists say, the high sensitivity of the CDI suggests that CDI-like tests could also prove useful for diagnosing other neurodegenerative diseases, such as Alzheimer's disease, Parkinsons's disease and fronto-temporal dementias, all of which, like prion diseases, involve various forms of protein misprocessing. These diseases currently are diagnosed by neuropathological analysis and immunohistochemistry.
"Whether immunohistochemistry underestimates the incidence of one or more of these common neurodegenerative diseases is unknown, but the CDI could shed light on these diseases," says co-author Bruce Miller, MD, UCSF A.W. and Mary Margaret Clausen Distinguished Professor of Neurology and director of the UCSF Memory and Aging Center.
The finding will be printed on-line and in print on March 1, 2005 in Proceedings of the National Academy of Sciences.