Development of a fully automated sample-to-answer system for rapid molecular detection of pathogens in whole blood.

BACKGROUND

Sepsis is a life-threatening organ dysfunction caused by infection and remains a major cause of mortality worldwide. Clinical outcomes depend on rapid pathogen identification and timely initiation of targeted antimicrobial therapy. Conventional blood culture requires 24-72 h, and although qPCR shortens turnaround time, conventional workflows still depend on labor-intensive sample preparation and open handling steps. An automated diagnostic workflow that can directly process whole blood from collection tubes is therefore needed. The problem addressed in this study is the lack of a rapid, fully automated sample-to-answer platform for sepsis pathogen detection from whole blood.

RESULTS

We developed a fully automated sample-to-answer platform that integrates robotic tube handling, including automated decapping and aliquoting, magnetic bead-based nucleic acid extraction, an ultrafast real-time PCR module, and a compact CMOS-based fluorescence detector. The system was evaluated using phosphate-buffered saline (PBS) and human whole blood spiked with Salmonella spp. DNA purified by the automated workflow showed purity comparable to that obtained by manual extraction with a commercial kit, confirming biochemical reliability. The complete process, from DNA extraction to 40 PCR cycles, was finished within 30 min. The limits of detection were 1 × 102 CFU/mL in PBS and whole blood, matching the performance of commercial benchtop qPCR instruments.

SIGNIFICANCE: This study demonstrates a closed, fully automated sample-to-answer system that performs direct whole-blood processing without manual intervention. Its novelty lies in integrating robotic handling, on-cartridge extraction, rapid amplification, and compact fluorescence detection into a single workflow. By reducing hands-on steps and contamination risk while maintaining sensitivity and speed, the platform offers a practical route toward point-of-care sepsis diagnostics and earlier targeted treatment.