Diagnostic method for direct analysis of human material collected in vivo
The technology facilitates simplified and improved cancer diagnostics by combining cell analysis and in situ multiplex molecular profiling of nucleic acid and protein biomarkers on in vivo collected material, such as circulating tumor cells (CTCs).
This enables clinical decision making by monitoring disease progression and treatment response and opens up for future screening of risk groups.
Current assays for bulk analysis of a larger number of cells may not detect molecules in rare cells and do not provide information on the cellular origin of each molecule. Thus, it is desirable to detect mRNA in situ directly within single cells in a specific and quantitative manner within a heterogeneous population of cells. Additionally, single molecule sensitivity, ultimately down to single nucleotide resolution, is vital for studying sequence variation such as point mutations or splice variations of expressed transcripts. Single nucleotide sequence variants as well as splice variants have been identified as important diagnostic disease biomarkers, particularly for diagnosis of different cancers.
The technology enables localized multiplex detection of RNA or proteins in situ within circulating cells or free circulating nucleic acids captured in vivo by a collection device. It comprises a functional, three-dimensional capture surface equipped with detection receptors. The molecular assays are based on highly specific padlock probe ligation or proximity ligation assay (PLA) followed by rolling circle amplification (RCA).
Quantification may occur directly on the device by suitable labelling methods. In combination with next generation barcode sequencing, a highly multiplex analysis is possible. Thus, in situ resolving of sequence variation and determination of transcriptional expression at single cell level in heterogeneous cell populations is possible. Additionally, it may be combined with conventional cell staining assays, providing information on cell morphology.
The technology combines cellular and molecular information of a biomarker enabling very specific diagnostics.
- Very versatile and broadly applicable, but also focused
- Enrichment of different types of human material
- Potentially higher yield than other technologies
- Significantly less human material necessary
- Direct visualization of sequence variations
Wissenstransferzentrum Süd (WTZ Süd)
Technische Universität Graz Graz University of Technology, Mandellstraße 9/II
StichworteCELL PROFILING, FREE CIRCULATING, NUCLEIC ACIDS, PROTEINS, CIRCULATING TUMOR CELLS (CTC), PADLOCK PROBE