Multiplexed Detection of Biomarkers in Lateral-Flow Immunoassays

Multiplexed detection of biomarkers, i.e., simultaneous detection of multiple biomarkers in a single assay, is a process of great advantages including enhanced diagnostic precision, improved diagnostic efficiency, reduced diagnostic cost, and alleviated pain of patients. A typical lateral-flow immunoassay (LFIA) is a widely used paper-based immunochromatographic test strip designed to detect a target biomarker through two common formats: sandwich assay and competitive assay. In order to obtain qualitative or quantitative result, a probe with unique optical or magnetic property is usually employed to characterize the concentration of the target biomarker. The typical LIFA is suitable for point-of-care testing due to its simplicity, portability, cost-effectiveness, and rapid detection of a target biomarker. However, detection of a single biomarker in the typical LFIA is not favorable for high throughput. Therefore, multiplexed detection of biomarkers in LFIAs has been extensively studied in recent years for high throughput. To accomplish multiplexed detection of biomarkers in LFIAs, the most frequently used method is to increase the number of test lines (TLs) on a nitrocellulose (NC) membrane. Alternatively, integration of multiple test strips together, where each test strip has one TL on the NC membrane, is employed for multiplexed detection of biomarkers. Sometimes, a single test strip with only one TL on the NC membrane is applied for multiplexed detection of biomarkers. This paper reviews three common structures for multiplexed detection of biomarkers in LFIAs, i.e., a test strip with multiple TLs, a multi-channel structure with multiple test strips, and a test strip with a single TL. Based on the optical and magnetic properties of probes, different signal detection strategies that include colorimetric detection, fluorescent detection, surface-enhanced Raman scattering detection, and magnetic detection, along with performance and perspectives are discussed.