Development of a semi-quantitative lateral flow immunoassay to classify the levels of SARS-CoV-2 antibodies in plasma

Lateral flow immunoassays (LFIAs) are versatile paper-based detection devices widely used for rapid on-site biomarker detection, yet they remain largely qualitative, which limits their utility in point-of-care (POC) settings. This study presents a colorimetric, semi-quantitative, and equipment-free LFIA format using a barcode-style design to categorize antibody levels as low, medium, or high. Using anti-SARS-CoV-2 antibodies in human plasma as a model, a multi-line assay was constructed with highly spherical gold nanoparticles (AuNPs) for improved sensitivity. The barcode-style LFIA demonstrated strong agreement with the validation method, with 90 % of results falling within ±1 category of the reference values. Additionally, the impact of AuNP morphology was investigated by comparing the assay performance with highly spherical (S), commercial (C), and quasi-spherical (Q) nanoparticles. While S-AuNPs improved the detection limit by two-fold in spiked plasma samples, all AuNP types performed comparably in clinical plasma. These results demonstrate a promising strategy for bridging the gap between qualitative and quantitative testing in low-resource or POC settings, enabling more informative readouts without the need for instrumentation.