Ultra-sensitive and high efficiency detection of multiple non-small cell lung cancer-related miRNAs on a single test line in catalytic hairpin assembly-based SERS-LFA strip

Accurate quantification of multiple miRNAs biomarkers in body fluid is still a challenge for early screening of cancer. Herein, by catalytic hairpin assembly as a signal amplification strategy, we designed a novel surface-enhanced Raman scattering (SERS)-lateral flow assay (LFA) strip for ultrasensitive detection of miR-21 and miR-196a-5p in non-small cell lung cancer (NSCLC) urine on a single test (T) line. 4-mercaptobenzoic acid or 5,50-dithiobis-2-nitrobenzoic acid as Raman molecules was labeled and two hairpin DNA sequence was modified gold nanocages (GNCs) were designed as two SERS tags. Through target miRNA-triggered catalytic hairpin assembly (CHA), the double-stranded DNAs (H1-H2 complex) formed by SERS tags and the related hairpin-structured DNA sequence 2 (H2) were immobilized on a single T line of SERS-LFA strip. This generated abundant "hot spots" because of the formation of numerous H1-H2 complex thus facilitated the SERS measurement. Through this method, two kinds of miRNAs were analyzed, resulting in limits of detection of 2.08 pM and 3.31 pM for miR-21 in PBS buffer and human urine, 1.77 pM and 2.18 pM for miR-196a-5p in PBS buffer and human urine. Significantly, the SERS-LFA strip exhibited high specificity and good repeatability toward miRNAs. The whole detection