Development of highly sensitive and specific methods for early detection of protein aggregation diseases; Proximity Ligation-based detection of prion diseases
2007: 2 years / Grant: 40 000 euros
Project: The “PrioDia” project proposed to develop a novel method of detection for abnormal aggregated proteins and peptides involved in some neurodegerative diseases. This method was based on the aggregation properties of these abnormal peptides and proteins, the use of specific antibodies and on the «solid phase proximityligation assay: SP-PLA»that allowed the co-locatization of 3 or more copies of a specific protein or peptide.
Results: For prion disorders, the group developed the SP-PLA method for the detection
of PrP aggregatesin brain homogenatesfrom infected hamsters(107 fold diluted). The SP-PLA-based
detection of aggregated PrP was demonstrated using either of the 2 monoclonal anti-PrP antibodies,
3F4 and 6H4.
For Alzheimer’s disease, a specific detection of A protofibrils was obtained in SP-PLA with the mAb158 monoclonal antibody, and the team was able to detect 0.1pg/ml A protofibrils. The assay was also used to detect soluble A aggregates in brain homogenates from an Alzheimer transgenic mice model.
Another format of the technology, in situ PLA, was utilized to studythe microtubule-affinityregulating kinase (MARK) mediated phosphorylation of Tau at Ser262 site in Alzheimer’s disease using transfected cells and human brain tissue sections. The study demonstrated, for the first time, that MARK4 isthe predominant isoform of the MARK familyto mediate the pathological phosphorylation of tau