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Novel Aggregated Biomarker Assay Technology

The challenge in developing blood tests for these aggregated proteins is to achieve the required level of sensitivity (lowest practical detection limit) and appropriate specificity for the oligomeric and aggregated forms.

CynapseDx has incorporated three technical enhancements to allow us to measure the oligomeric and aggregated forms of these biomarker proteins in blood.

  1. Immunoconcentration.

    We use antibody-coated magnetic beads to scavenge proteins from a whole blood sample. CynapseDx uses large (50-100 µm diameter) dense beads with a large, dense magnetic core. This product was originally developed by us to harness cells from blood or homogenised tissues and is ideal for navigating within the whole blood matrix. The dense beads travel readily throughout the matrix until they encounter the target protein. The beads are coated with antibodies against ß-amyloid, α-synuclein or phospho-tau, which bind to the appropriate protein whether it is free in solution, complexed with other proteins, bound to intact cells or associated with cell membrane fragments.

    The beads capture protein from a large blood volume and after washing the antibody interaction is reversed and the target protein is eluted into a small volume, increasing the concentration by a factor >10x for subsequent analysis

  2. Immobilisation of aggregated proteins.

    The expertise in measurement of aggregated proteins developed in the late 1990’s, when there was an urgent need to detect the abnormal aggregated form of prion protein in the brains of cattle affected with Bovine Spongiform Encephalopathy (BSE or mad cow disease). CynapseDx has adapted the market leading BSE testing product for measuring the oligomeric and aggregated forms of brain-derived proteins for the current neurodegenerative disease application. The BSE detection kits are based on a microplate format where the wells are coated with a mixed mode charged polymer (Seprion) that, under selective conditions, exhibits specificity for the oligomeric and aggregated forms of a protein over the monomeric form.  The surface-bound Seprion ligands used in the BSE kit are generic for binding aggregate forms of all proteins and so do not display specificity for a particular protein biomarker.

  3. Ultrasensitive labels.

    The final step in the CynapseDx assay procedure uses a labelled antibody conjugate that binds to the aggregated protein immobilised to the Seprion ligand on the microwell surface.

    In this immunometric, or sandwich assay format a very high sensitivity is achieved by the use of an ultrasensitive label.

    The combination of all 3 enhancements allows for the highly sensitive and specific detection of the oligomeric and aggregated forms of each biomarker protein in blood.