Abstract
Objectives: Multiple Sclerosis is a chronic, enigmatic, progressive central nervous system “inflammatory” disease, with no know etiology. As with other “inflammatory” diseases there is the possibility that a cryptic infectious trigger may play a role in initiating Multiple Sclerosis. M. avium subspecies paratuberculosis causes Johne’s disease in ruminants and may be an infectious trigger in Crohn’s disease.
In this study, frozen archived brains from patients with Multiple Sclerosis, pure culture of multiple bacteria and circulating WBCs were assayed with proprietary (AffymetrixÔ RNA view®) Tissue and Cell fluorescent in situ hybridization assay for MAP RNA.
Results: Repetitively, false positive signal was observed in the “No-Probe” negative control. Despite advice from the technical staff at Affymetrix, multiple experimental modifications could not prevent positive signal in the “No-Probe” negative control.
When studying human white blood cells under specific storage conditions, we observe positive signal with human house-keeping genes, when no signal is seen in the No-Probe controls.
Conclusions: We conclude, that when performed according to manufactures instructions and with multiple variations on the manufactures recommended suggestions to correct for false positive signal, that the AffymetrixÔ RNA view® TISSUE assay cannot be used to detect M. avium subspecies paratuberculosis in pre-frozen brains of humans with Multiple Sclerosis. In contrast, using the AffymetrixÔ RNA view Cell fluorescent in situ hybridization system, evaluating human white blood cells, we reliably identify human house-keeping genes. This indicates that the Cell fluorescent in situ hybridization assay may be useful when evaluating circulating cells for specific pathogens.