Process parameter optimization for lateral flow immunosensing

Abstract Lateral flow immunoassay (LFIA) is a widely used technique in the field of point-of-care diagnostics. Assembly of components such as sample pad, conjugate pad, nitrocellulose membrane (NCM), absorbent pad on the plastic backing laminate to prepare the LFIA strip for analysis is an important factor. Processing of the strip assembly and heat or buffer treatment of strip affects the surface morphology, structure and porosity of the NCM. Flow time and pattern of the fluid flow through the LFIA strip surface affects the sensitivity of the LFIA. In this work, we have studied various combinations of strip component assemblies, the morphology of NCM, and flow time and flow pattern of fluid through the strip surface. The LFIA is developed using the optimized strip assembly for the detection of rabbit IgG, which is commonly used as the substrate for control line formation in most of the LFIA tests. Assembly of the sample pad overlapping on the conjugate pad that overlaps on the NCM at one end of the strip and absorbent pad overlapping on the NCM from other end was found to be optimal. Uniform, leveled, slower fluid flow was observed with this assembly when dried at 55 °C for 20 min. Overlap of the membranes is critical for wicking as the fluid flow through membrane because of capillary force. An optimal concentration of the rabbit IgG required for the control line was found to be 1.2 µg/mL. The reported methodology may be advantageous for the development of LFIA for detection of various biomarkers with improved sensitivity.