Enabling Multi-Analyte Analysis of Lung Cancer FFPE Samples with 3D Biology™ Technology
Worldwide, lung cancer is the most commonly diagnosed form of cancer with a survival rate among the lowest. Somatic mutations (in the form of SNVs and InDels) and gene fusions account for the majority of clinically interpretable and actionable genomic alterations. Biomarkers for lung cancer and identification of molecular drug targets are also actively sought through protein analysis methodologies. Multi-analyte analysis of DNA, RNA, and protein is often hindered by limited amounts of FFPE-preserved specimens. When enough sample is available, each analyte is typically profiled on different instrument platforms, which requires complex workflows and specialized data analysis expertise. To provide a unified and simpler alternative, NanoString’s molecular barcoding technology has been adapted to permit simultaneous digital measurement of biologically-relevant targets that span DNA, RNA, and protein, known as 3D Biology™ technology. This approach has several advantages over other analytical methods. Direct, single-molecule digital counting allows detection over a broad dynamic range with high reproducibility, often >98% concordance between technical replicates. SNV probes enable sensitive and specific identification of ≤ 5% DNA mutant allele sequences from 5 ng of FFPE-extracted genomic DNA. Fusion transcripts are detected with 5’/3’ imbalance probes and toehold-mediated junction probes. Protein and phospho-protein expression is measured via DNA-labeled antibodies and a single antigen retrieval step. The 3D Biology workflow requires just two 5-10 µM sections of FFPE tissue; RNA/DNA extracted from one section and multiplex digital protein profiling from the second.