Abstract Comprehensive genomic profiling (CGP) guides precision oncology, yet access remains limited in community and regional laboratories. SeqF is a Nanopore-based targeted sequencing workflow for low-throughput assays, utilizing the MinION™ device with standard molecular biology equipment and dedicated bioinformatic analysis software. SeqF was evaluated using 40 DNA samples, derived from clinical and reference specimens, with 20 samples analyzed in each of two targeted panels: EGFR-pathway (commercial off-the-shelf amplicon panel; 12 amplicons across 4 genes; ∼1.5 kb total) and MPN (custom amplicon panel; 80 amplicons across 20 genes; ∼35 kb total). Testing was performed in a clinical research and development laboratory. The wet-lab workflow comprises PCR-based target enrichment, sample indexing/barcoding, and incorporation of unique molecular identifiers (UMIs) for the EGFR panel only. Indexed libraries then undergo rolling-circle amplification followed by adapter ligation for Nanopore sequencing. Library preparation spans two days, followed by ∼12-24 hours of sequencing (runs may be stopped early once coverage targets are met). The analysis pipeline is run via a user interface and involves base calling from raw sequencing reads (HAC in MinKNOW), followed by an automated pipeline that includes demultiplexing, consensus (error correction), alignment, variant calling, annotation, and QC. When compared to sequencing performed on Illumina instruments, and using paired tumor specimens, SeqF achieved 99.7% concordance for single-nucleotide variants at ≥5% variant allele frequency (VAF). Correlations between the measured VAFs (5-40%) in clinical samples and reference standards were ≥95% in both panels tested. The workflow demonstrated feasibility for small batch sizes and delivered robust analytical performance. The process was completed in under 72 hours, from extracted sample to VCF file and sequencing run QC report generation. SeqF enables decentralized tumor profiling with low infrastructure requirements and rapid results, potentially expanding access to precision oncology in community laboratory settings. The streamlined workflow and analysis pipeline may help democratize access to molecular diagnostics, ultimately improving patient outcomes through more timely and personalized treatment decisions. Future enhancements such as expanded panel content, broader UMI integration, and real-time variant interpretation will significantly increase the platform’s utility. This increased utility will enable scalable precision oncology across a wider range of cancer types and healthcare environments. Citation Format: Kimberly A. Holden, Roi Feingersch, Dvir Dahary, Margalit Feiger, Tal Havkin-Solomon, Benjamin M. Cohen, George Way, Shakti Ramkissoon, Marcia Eisenberg, Brian Caveney, Eric Severson, Taylor J. Jensen, Jonathan Williams. SeqF™: evaluation of an affordable nanopore-based assay for decentralized tumor profiling abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 3246.
Holden et al. (Fri,) studied this question.