A functionalized Sup35NM nanofibril-assisted oriented antibody capture in lateral flow immunoassay for sensitive...

Rapid and sensitive dengue non-structural protein 1 (NS1) detection assay is essential for the treatment of disease and currently releases high medical cost burdens. To address the limitations of conventional LFIA strips, we have developed an improved Sup35NM-Z-based LFIA that immobilizes antibodies on cellulose membranes in an orientated manner to increase the sensitivity of LFIA strips. A dual-functional Sup35NM nanofibril was fabricated by fusion with the antibody binding domain; resultant nanofibril from the amyloid Sup35NM was sprayed on the T-line to orientate the capture antibody and produces fluorescence signals. Antibody binding analysis showed that self-assembly of the Sup35NM monomer does not affect the binding activity of the Z-domain with the antibody. The NS1 for DENV-2 infection was chosen as a model target antigen to assess the feasibility of the Sup35NM-Z-domain-based LFIA platform. Under optimal conditions, the Sup35NM-Z-domain-based LFIA detected NS1 within 15 min with a detection limit of 1.29 ng/ml, while the detection limit of traditional LFIA with the same concentration of anti-NS1-Ab1 on the T-line by conventional physical adsorption was 2.20 ng/ml, 1.7 times higher than that of Sup35NM-Z-domain-based LFIA. As compared to traditional LFIAs, the Sup35NM-Z-based LFIA had a wide detection range of 1.29-625 ng/mL. The LFIA's clinical performance in identifying NS1 was also assessed using 15 clinical samples. The LFIA accurately recognized positive and negative samples, equal to 86.7% accuracy. The developed Sup35NM-Z-domain-based LFIA in this study offers great potential for the identification of target markers because of its greatly improved sensitivity and wider detection range.