MOLECULAR TECHNIQUES GUIDING ASSAY DESIGN | qPCR
Choices in Molecular Techniques Guide Assay Design
MUTATION DETECTION & GENOMIC COVERAGE DIFFER BY SEQUENCING METHOD
For assay developers, meeting both clinical and operational requirements requires balancing analytical performance, cost-efficiency, and usability. Precision oncology depends on technologies that accurately detect genetic alterations, guide treatment decisions, and deliver results that can be effectively interpreted across diverse healthcare settings. In today’s commercial landscape, three techniques— qPCR , targeted next-generation sequencing ( tNGS ), and whole-genome sequencing ( WGS )—dominate. These techniques underpin most regulatory-approved oncology assays, owing to their clinical utility, scalability, and established regulatory pathways. 10, 11 Selecting the right technology requires careful alignment with the assay’s intended use, as each one presents trade-offs in turnaround time, assay breadth, infrastructure demands, and accessibility. These factors must be weighed during assay design and commercialization to maximize both clinical utility and market viability.
qPCR Precision with Speed and Simplicity Quantitative PCR (qPCR) is the most established and widely adopted molecular diagnostic technology for oncology. It forms the foundation of many FDA- approved companion diagnostics (CDx) (refer to Table 1 ), including therapies targeting EGFR mutations in non-small cell lung cancer (NSCLC), BRAF V600E in melanoma, 12 and KRAS wild-type status in colorectal cancer. 13 These assays typically use allele-specific amplification or probe-based detection methods to achieve precise, reproducible results with minimal input material.
qPCR’s primary strengths include fast turnaround time—often within hours—low cost per test, and high sensitivity for detecting single or limited mutations, making it ideal for cancers with well-characterized genetic drivers. However, its utility is limited to pre- defined targets and lacks the ability to detect novel or complex genomic alterations such as gene fusions or structural variants. While it remains a gold standard for detecting known driver mutations, qPCR’s narrower analytical range makes it less suitable for applications requiring broad genomic profiling or the detection of emerging biomarkers. 14
Table 1 . Examples of FDA-Cleared qPCR Precision Oncology Assays Used in Clinical Practice
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