BACKGROUND: Liquid biopsy is reshaping the landscape of cancer diagnostics by turning a simple blood sample into a rich source of real-time molecular insights. Among its most promising targets are circulating tumor cells (CTCs) and extracellular vesicles (EVs), which carry valuable clues about tumor progression, metastasis, and treatment response. However, traditional workflows for analyzing CTCs and EVs typically rely on immunoaffinity-based enrichment followed by molecular assays such as quantitative Polymerase Chain Reaction (qPCR) or Enzyme-Linked Immunosorbent Assay (ELISA). These approaches are often limited by low sensitivity, high costs, and complex procedures, hindering their widespread clinical use.
RESULTS: Recent advances in Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas technologies offer a multifaceted approach to biomarker analysis. Cas9 is primarily used for functional gene interrogation to identify and validate targets, while Cas12a and Cas13a serve as direct diagnostic tools, enabling ultrasensitive signal amplification for DNA, RNA, and protein markers. By integrating these distinct Cas effectors with aptamer-based recognition, nanomaterial-assisted enrichment, and hybrid amplification techniques like Hybridization Chain Reaction (HCR), Rolling Circle Amplification (RCA), researchers have developed highly sensitive and programmable platforms for analyzing CTCs and EVs.
SIGNIFICANCE: Despite ongoing challenges such as off-target effects, Protospacer Adjacent Motif (PAM) sequence limitations, and clinical variability, the field is rapidly evolving. The convergence of CRISPR diagnostics with artificial intelligence, device miniaturization, and multiplexed sensing is accelerating clinical translation. Collectively, these innovations are paving the way for a new era of precision oncology that is fast, flexible, and achievable with only a drop of blood.
