Protein misfolding

The misfolding of proteins and their subsequent aggregation in the brain is thought to be the cause of a group of related neurological diseases including Parkinson’s, Alzheimer’s and the transmissible spongiform encephalopathies such as BSE, scrapie, vCJD and CWD (the ‘protein conformational disorders’, or ‘protein aggregation diseases’). The development of highly sensitive and specific diagnostic assays for these diseases is a high priority.

The majority of assay kits for BSE and scrapie in brain tissue seek to differentiate normal prion protein (PrPsen) from the abnormal, aggregated form (PrPres) with a proteinase K digestion step. As an alternative approach Microsens has developed polymeric ligands (the ‘Seprion’ range) with specificity for PrPres. A group of polymeric compounds have been identified that have the ability to bind PrPres, under selective conditions, in BSE, scrapie, CWD and vCJD-infected tissues. It has also been shown that Seprion ligands can bind a PrPsc-like material (the scrapie form of PrPres) in infected sheep blood and a signal was also detected in the blood of a human patient with prion disease.

An immunometric assay format has been developed, with the Seprion ligands immobilised directly to the wells of a microplate. After addition of tissue homogenate (such as brain, lymph node or tonsil) and direct capture of PrPres the bound prion protein is detected with a specific prion protein-specific antibody/peroxidase conjugate. This assay procedure is greatly simplified compared to the proteinase K digestion methods and is suitable for automation on an ELISA processor.

Assay kits incorporating the Seprion ligands in the microplate format are now available from Idexx Laboratories Inc, Westbrook, USA for the detection of PrPres in brain tissue from BSE and scrapie-infected cattle and sheep and in brain and lymphoid tissue from CWD-infected deer. In addition Microsens has developed a microplate-based assay kit for the detection of PrPres in tonsil tissue from subjects infected with vCJD.

Microsens has also demonstrated that the Seprion ligand technology is applicable to the detection of the aggregated proteins present in the brains of Alzheimer’s Disease patients (amyloid and tau proteins) and Parkinson’s Disease (alpha synuclein protein) (unpublished results). This has recently been extended to the detection of aggregated huntingtin protein in tissues from a mouse model of Huntington’s Disease (unpublished results).

The Seprion ligands can also be coated onto the surface of magnetic beads as an alternative format to the microplate. Microsens is developing a general separation kit based on the microparticle format with applicability to the separation of all amyloid-type proteins.