Lateral flow assay with automatic signal amplification based on delayed substrate release.

The low sensitivity of Lateral flow assay (LFA) limits its application in rapid detection for trace targets. LFAs with nanozyme (nanozyme-LFA) as signal labels have demonstrated excellent performance in point of care testing (POCT). However, additional operational steps for substrate catalysis in nanozyme LFA are required, which makes the nanozyme-LFA operation complicated. In this work, we designed a LFA based on delayed substrate release (SGF-LFA), in which a commercialized glass fiber membrane embedded with substrate (SGF) was fixed at the sample pad. The SGF could automatically execute substrate delivery and catalysis, thus eventually achieving a one-step LFA operation for the nucleic acid detection of influenza A virus H1N1. In this SGF-LFA, 3,3 '- diaminobenzidine (DAB) was oxidized and deposited, producing a strong signal amplification under the catalysis of Au@PtNP nanozyme. The SGF-LFA could detect the nucleic acid of H1N1, with a linear range of 0.02-50 nM and a limit of detection (LOD) as low as 0.02 nM, which was 25-fold lower than that of the nanozyme-LFA before catalysis. In addition, the analytical performance was close to that of a manual operation mode of catalysis amplification. The application of SGF-LFA for detecting the H1N1 nucleic acid in serum samples obtained a recovery rate of 96 %-102.7 %, indicating that SGF-LFA has great potential in point-of-care testing.