Split CRISPR/Cas systems: Pioneering solutions for molecular diagnostics challenges

Area of Science:

• Molecular Biology
• Biotechnology
• Diagnostics

Background:

• Conventional CRISPR detection methods face limitations in sensitivity, specificity, and regulatory flexibility for low-abundance targets.
• Split CRISPR/Cas systems represent an advancement to overcome these challenges in molecular diagnostics.

Purpose of the Study:

• To review split-activation strategies in CRISPR/Cas systems for enhanced diagnostic performance.
• To highlight innovations in ultrasensitive RNA detection and multiplex nucleic acid analysis.

Main Methods:

• Focuses on split-activation strategies, including split activator-mediated Cas systems and split crRNA architectures.
• Examines applications in clinical and on-site monitoring.

Main Results:

• Achieves femtomolar-level sensitivity for biomarkers with superior single-base discrimination.
• Enhances analytical performance without requiring pre-amplification.

Conclusions:

• Split CRISPR/Cas systems offer significant improvements for precision diagnostics.
• Future directions involve customized Cas variants, nanomaterial-based workflows, and microfluidic platforms for broader applications, especially in resource-limited settings.