A SERRS-Based Lateral Flow Assay for Sensitive Detection of Mitochondrial DNA in Endometriosis Screening

Endometriosis (EMT) is an incurable and painful chronic illness that affects approximately 10% of people assigned female at birth worldwide. Currently, EMT takes on average 5-7 years to diagnose after histological confirmation with a tissue sample collected via laparoscopy. Therefore, there is a demand for developing new and powerful detection tools that can work in conjunction with imaging techniques to make EMT diagnosis more accessible and reduce diagnostic delays. This study proposes a proof-of-concept lateral flow assay (LFA) for the detection of an EMT biomarker based on mitochondrial DNA deletion mutations found in cell-free mitochondria in plasma. Surface-enhanced resonance Raman scattering (SERRS) was integrated with the LFA to increase sensitivity and allow quantitation using gold nanoparticles functionalised with the near-infrared fluorescent dye NIR 4f. The SERRS-LFA was compared to gel electrophoresis and colorimetry in a typical polymerase chain reaction (PCR) workflow. SERRS signals were analysed using confocal Raman microscopy in combination with digital SERRS. This digital SERRS-LFA demonstrated a limit of detection (LOD) of 5.6 × 10² input copies, about 109-fold lower than the reference colorimetric method. When mutant DNA was tested as abundance of background wild type DNA, an LOD of 0.35% was obtained, about 14-fold lower than the reference colorimetric method. Finally, quantitative analysis with a handheld Raman reader demonstrated the potential of bringing future amplification-free SERRS-based LFAs to point-of-care settings.