Questions & answers
Below are answers to some of the most pressing customer questions that were submitted during the webinar.
Liquid biopsy testing
Liquid biopsy relies on the dimension of the lesion. The smaller it is, the more difficult it will be to detect a mutation in the bloodstream. Locational imaging like HER2-PET still remains a useful diagnostic tool to assess the striations of the tumor.
dPCR and NGS technology
As shown during the presentation, we applied dPCR to track index mutations previously found by NGS analysis. In selected cases (e.g., in the case of a precise query from the oncologist), we also applied dPCR assays based on tissue findings without running a baseline NGS analysis.
Mutational insights
Yes, we also collected peripheral blood mononuclear cells (PBMCs) from the enrolled patients and tested these analytes for TP53 mutations. In two cases, aged 79 and 82, we confirmed that TP53 mutations seen as ctDNAs in the bloodstream were present as gDNA in the white cells, thus being derived from clonal hematopoiesis.
Sample collection
We monitored our patients by collecting blood every 21 days, at the occasion of chemotherapy infusion.
Sample extraction
We do recommend cfDNA or ctDNA/ctTNA isolation from body fluids, also including plasma, for different reasons, most of which are related both to research and diagnostic purposes (the latter within a molecular tumor board).
There are many kits on the market based on silica or magnetic beads. We routinely extract our ctTNAs from frozen plasma using a commercially available magnetic bead-based kit.
Data analysis
We routinely filter our NGS results by applying a preset filter in the analysis software. We also confirm every SNV found by NGS by running the specific dPCR assay. Only mutations that were called by the two platforms were considered "true" positives.